TRENDS R., Res, P.C.M. et al. (1991) J. Exp. Med. 174, 583-592 5 Yssel, H., Shanafelt, M-C., Soderberg, C. et al. (1991) J. Exp. Med. 174, 593-601 6 Wierenga, E.A., Snoek, M., de Groot, C. et al. (1990).1- Immunol. 144, 4651-4656 7 Parronchi, P., Macchia, D., Piccinni, M-P. et al. (1991) Proc. Natl Acad. Sci. USA 88, 4538-4542 8 Maggi, E., Del Prete, G-F., Macchia, D. et al. (1988) Eur. ]. lmmunol. 18, 1045-1050 9 Paliard, X., de Waal Malefit, R., Yssel, H. et al. (1988)J. ImmunoL 141,849-855 10 Swain, S.L., Weinberg, A.D. and English, M. (1990)J. lmmunol. 144, 1788-1799 11 Gaiewski, T.F., Joyce, J. and Fitch,
F.W, (1989) I. Immunol. 143, 15-22 12 Coffman, R.L., Chatelain, R., Leal, L.M.C.C. and Varkila, K. (1991) Res. Immunol. 142, 35-40 13 LeGros, G., Ben-Sasson, S.Z., Seder, R., Finkelman, F.D. and Paul, W.E. (1990) I. Exp. Med. 172, 921-930 14 Locksley, R.M., Heinzel, F.P., Holaday, B.G. et al. (19911 Res. ImmunoL 142, 28-31 15 Maggi, E., Parronchi, P., Manetti, R. et al. (1992) J. Immunol. 148, 2142-2147 16 DeMaeyer, E. and DeMaeyerGuignard, J. (1988) Interferons and Other Regulatory Cytokines, John Wiley & Sons 17 Parronchi, P., De Carli, M., Manetti, R. et al. J. Immunol. (in press)
18 Kobayashi, M., Fitz, L., Ryan, M. et al. (1989)J. Exp. Med. 170, 827-845 19 Finkelman, F.D., Svetic, A., Gresser, I. et al. (1991)I. Exp. Med. 174, 1179-1198 20 Paul, W.E. (1992) in New Advances on Cytokines (Romagnani, 5., Abbas, A.K. and Mossman, T.R., eds), pp. 151-154, Raven Press 21 Ben-Sasson, S.Z., LeGros, G,, Conrad, D.H., Finkelman, F.D. and Paul, W.E. (1990)Proc. Natl Acad. Sci. USA 87, 1421-1425 22 Piccinni, M-P., Macchia, D., Parronchi, P. et al. (1991) Proc. Natl Acad. Sci. USA 88, 8656-8660 23 Conrad, D.H., Ben Sasson, S.Z., LeGros, G., Finkelman, F.D. and Paul W.E. (1990) 1. Exp. Med. 171, 1497-1508
Immunotoxins: the power and the glory Edward J. Wawrzynczak and Elaine J. Derbyshire
The traditional triumvirate of natural toxins - diphtheria toxin (DT), Pseudomonas exotoxin A (PE) and ricin - all act by binding to cells, undergoing internalization and intracellular processing; this allows ddivery to the cytoso[ of a catalytic component capable of inactivating protein synthesis. The secret of making a useful immunotoxin is to eliminate the cellbinding ability of the toxin without loss of the other functions vital to its action. Determination of the crystal structure of DT, presented by S. Choe (National Institutes of Health, Bethesda) ~, revealed a Y-shaped molecule with clearly delineated domains involved in receptor binding, translocation and catalysis. The receptor-binding domain is a flattened 13-barrel reminiscent of the immunoglobulin (Ig) fold. It was suggested that complete replacement of the receptor-binding domain by an Ig variable domain (F0 could redirect the binding specificity of DT.
Immunotoxins are hybrid proteins in which the potent cytocidal action of a toxin is harnessed for the selective destruction of target cells by attachment to a specific monoclonal antibody (mAb) or growth factor. This brief article describes the latest advances in the molecular and cellular biology, pharmacology and clinical evaluation of immunotoxins, as discussed at a recent meeting.
Elucidation of the DT structure has helped to rationalize experimental findings with recombinant single-chain hybrid fusion proteins between DT analogues and growth factors produced in Escherichia coll. L-F. Jean, F. Lakkis (University Hospital, Boston) and J. Shaw (Seragen Inc., Hopkinton) reported The Third International Symposium on lmmunotoxins was organized by Arthur E. Frankel and was held at the Sheraton World Resort, Orlando, Florida, USA on 19-21 June 1992.
the properties of fusions between interleukin 4 (IL-4), 1L-6 and epidermal growth factor (EGF) with DAB389, a truncated DT analogue. DAB3s9 completely lacks the recepto>binding domain and has superior targeting properties compared with DAB4s6, from which only part of the receptor-binding domain has been excised 2. The complexity of the mechanism of action of PE has been revealed in a series of elegant experiments by D. FitzGerald, I. Pastan and colleagues (National Institutes of Health, Bethesda). PE binds to the ubiquitous c~macroglobulin receptor. Following endocytosis, the toxin is cleaved after Arg279 by a cellular protease, to generate a cell-binding fragment and a catalytic fragment which becomes translocated to the cytosoP. Trp281 appears to be essential to an interaction with an intracellular component after the cleavage step. Gly280, the amino terminus of the catalytic fragment,
1992, Elsevier Science Publishers lid, UK.
Immunology Today
381
vo~. ~3 No. 1019v2
TRENDS
can be changed to Met280 without influencing translocation, allowing the creation of novel PE analogues no longer requiring intracellular proteolysis, a potential variable in the sensitivity of cells to immunotoxin. C. Siegall (Bristol-Myers Squibb, Wallingford) and U. Brinkmann (National Institutes of Health, Bethesda) described preclinical studies of recombinant single chain Fv fusions with truncated forms of PE lacking the cell binding domain, BR96(single-chain Fv)-PE40 and B3(single-chain Fv)-PE38KDEL. Both immunotoxins induced complete remission of large solid subcutaneous tumour xenografts in nude mice. The recombinant immunotoxins were superior in vivo to immunotoxin analogues made by chemical conjugation because of higher potency, lower toxicity, and better turnout penetration4. The attachment of high-affinity ligands to the two galactosebinding sites of ricin results in a blocked ricin with 1 000-fold lower cell binding, but there is evidence that the residual binding capacity is essential for the activity of blocked ricin immunotoxinss. R. Youle (National Institutes of Health, Bethesda) reported collaborative studies with M. Lord (University of Warwick) using recombinant ricin molecules in which either one or both binding sites had been incapacitated by site-directed mutagenesis. Wild-type and mutant recombinant ricin B (binding) chains were expressed in Xenopus oocytes and reassociated with recombinant ricin A (active) chain. The mannosyl side chains of the B chain mediated uptake of the reconstituted toxins by macrophages. Removal of both binding sites abolished toxicity, indicating the importance of galactose-binding in efficient delivery of the A chain to the cytosol6. The A chain of ricin can itself be targeted by direct attachment to mAb via a disulphide bond. A variety of other ribosome-inactivating proteins (RIPs) from plants and fungi, including gelonin, saporin, trichosanthin and mitogillin, have been cloned and expressed in E. coli. S. Bernhard (XOMA Corp.,
B-cell lymphoma using an antiCD19 immunotoxin, HD37-deglycosylated ricin A. In a severe combined immunodeficiency (SCID) mouse model of disseminated Bcell lymphoma developed by M-A. Ghetie (University of Texas, Dallas) 9, a mixture of the antiCD22 immunotoxin and either the anti-CD19 immunotoxin or the CD19 mAb gave superior antitumour effects, providing a rationale for combination therapy. Clinical trials Immunotoxins made with natuH65-ricin A chain (now known under the trade name 'CD5 Plus'), ral RIPs have recently reached the an immunotoxin that was directed stage of clinical evaluation. B43antiviral against the CD5 antigen, has (anti-CD 19)-pokeweed been administered to hundreds of protein, developed by F. Uckun patients with graft-versus-host dis- and colleagues (University of ease (GVHD) following allogeneic Minnesota, Minneapolis), demonbone marrow transplantation. P. strated significant anti-tumour Scannon (XOMA Corp., Berkeley) effects in a SCID mouse model, reported that when two groups especially in combination with cyof patients with steroid-resistant clophosphamide ~°, and has recently GVHD were treated with the entered phase I trials. B. Falini immunotoxin, enhanced resol- (University of Perugia, Italy) reution of acute GVHD occurred and ported early results of trials in there was a survival benefit in patients with refractory Hodgkin's grade I-III GVHD compared with disease treated with an anti-CD30 historical controls. Studies are in immunotoxin, Ber-H2-saporin J~. progress with 'CD5 Plus' in F. LeMaistre (University of patients with rheumatoid arthritis, Texas, San Antonio) and T. type I diabetes mellitus and other Woodworth (Seragen Inc., Hopkinautoimmune diseases. ton) reported trials with the first M. Grossbard (Dana-Farber recombinant immunotoxin in the Cancer Institute, Boston) pre- clinic, DAB486IL-2. This immunosented the results of a number of toxin selectively eliminates tumour trials using anti-B4-blocked ricin, cells and activated T cells that directed against the CD19 pan-B- express the high affinity IL-2 cell antigen, in patients with B-cell receptor and is thus applicable to a neoplasms refractory to conven- variety of haematological maligtional treatments7. A phase I trial nancies 12,~3, rheumatoid arthritis of adjuvant therapy for patients and insulin-dependent diabetes with non-Hodgkin's lymphoma mellitus. Future trials are planned following autologous bone marrow with another recombinant immunotransplantation is in progress. toxin, DAB3s9IL-2. E. Vitetta (University of Texas, Although many trials are still at Dallas) presented the results of a a preliminary stage, it is possible to collaborative study among several draw some general conclusions. centres comparing the behaviour of First, all immunotoxins show a two deglycosylated ricin A chain similar (though not identical) patimmunotoxins linked either to the tern of transient and reversible anti-CD22 mAb RFB4 or its Fab' toxicities; primarily, an elevation fragment, in phase I trials in of liver transaminases and/or a patients with B-cell lymphoma8. vascular leak syndrome characterDespite differences in potency, ized by hypoalbuminaemia and pharmacokinetics, toxicity and im- oedema. Secondly, a number of munogenicity, the overall perform- long-lasting remissions were ance of both agents was similar. achieved in dose-escalation studies M. Stone (Baylor-Sammons Cancer and anti-tumour effects were seen Center, Dallas) reported prelimi- in heavily pretreated patients. nary data of a new phase I trial in Thirdly, host antibody responses to Berkeley) used mutagenesis of recombinant single chain RIPs to create analogues with a single, free thiol group for site-specific conjugation to mAb. The position of the thiol on the RIP influenced enzymic activity, conjugation efficiency and immunotoxin potency; some RIP analogues formed more potent immunotoxins than the natural RIP or ricin A chain.
Immunology Today
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Vol 13 No. 10 •992
TRENDS
immunotoxins occurred in many patients, even in a proportion of immunosuppressed lymphoma patients.
mAbs that recognize human antigens associated with tumour neovasculature, which could be applied to many different types of tumour. Although much of the focus of Novel approaches immunotoxin research is directed The immune response to murine towards alleviation of human lg and toxin molecules of bacterial disease, immunotoxins are also or plant origin limits the amount of proving to be useful experimental immunotoxin that can be admin- tools. R. Wile}' (DVAMC and istered to patients. D. Newton Vanderbilt University, Nashville) (National Institutes of Health, described the use of immunotoxins Bethesda) found that bovine pan- to ablate specific neurones in excreatic RNAse linked to an anti- perimental animals. A saporin transferrin receptor mAb displayed immunotoxin directed against the selective cytotoxic activity against rat low-affinity neurotrophin receptumour cells in tissue culture L4. tor ~s was highly effective in Despite considerably lower potency destroying peripheral neurones than an analogous ricin A chain when administered systemically to rats. Microimmunotoxin, the RNAse immuno- Sprague-Dawley toxin exerted similar growth in- injection into the anterior lateral hibitory effects when injected into ventricle selectively destroyed neuglioblastoma xenografts. RNAses rones of the cholinergic basal foreof mammalian or human origin are brain. Since these are the neurones likely to be much less immunogenic that predominantly degenerate in than the conventional toxins, es- Alzheimer's disease, this rat model pecially in combination with may be helpful in investigating the pathophysiology of the disease and human or humanized mAbs. Conventional immunotoxins in testing potential therapies. readily reach turnout cells close to blood capillaries but penetrate Conclusion The rapid advances in our undersolid tumour masses poorly. A complementary approach to killing standing of toxin structure and malignant cells directly is to target action, new techniques for molecuthe vasculature upon which the lar engineering of immunotoxins, tumour relies for its nutrient promising results of recent patient supply. P. Thorpe and F. Burrows studies, novel approaches, and (University of Texas, Dallas) expanding interest suggest that a presented a mouse model of vascu- glorious future lies ahead for the new generation of lar targeting. The neuroblastoma powerful cell line C1300, transfected with immunotoxins. the gamma-interferon (IFN-y) gene, was grown in BALB/c nude mice. Eddie Wawrzynczak is at the IFN-y secreted by the tumour Institute o f Cancer Research, induced the expression of major Sutton, Surrey, UK SM2 5NG; histocompatibility complex (MHC) Elaine Derbyshire is at the class II antigens on the endothelial University o f Texas Southwestern cells adjacent to the turnout. An Medical Center, Dallas, T X 75235anti-class I1 immunotoxin elicited 8576, USA. extensive turnout necrosis through a process of endothelial cell damage, platelet deposition and References intravascular thrombosis; only a 1 Choe, S., Bennett, M.J,, Fujii, G. thin rind of turnout cells adjacent et aL (1992) Nature 357, 216-222 to normal skin blood vessels 2 Shaw, J.P., Akiyoshi, D.E., Arrigo, survived the immunotoxin treat- D.A. et al. (1991).I. Biol. Chem. 266, ment. When the treatment included 21118-21124 3 Fryling, C., Ogata, M. and a second immunotoxin directed FitzGerald, D. (1992) Infect. lmmun. against the tumour itself, the 60,497-502 readily accessible cells were also 4 Brinkmann, U., Pai, L.H., killed and complete turnout re- FitzGerald, D.J. et al. (1991) Proc. Natl gressions resulted. The hunt is on for Acad. Sci. USA 88, 8616-8620
t,,,,,,,ology r,,aa,,
383
5 Goldmacher, V.S., Lambert, J.M. and Blattler, W.A. (I 992) Biochem. Biophys. Res. Commun. 183~ 758-766 6 Newton, D.I,., Wales, R., Richardson, P.T. et al. (1992)1. Biol. Chem. 267, 11917-11922 7 Grossbard, M.L., Freedman, A.S., Ritz, J. et al. (1992) Blood 79, 576-585, 8 Vitetta, E.S., Stone, M., Amlot, P. et al. (1991) Cancer Res. 5 I, 4052-4058 9 Ghetie, M-A., Richardson, J., Tucker, T. et al. (1991) Cancer Res. 51, 5876-5880
10 Uckun, F.M., Chelstrom, L,M., Finnegan, D. et al. (1992) Blood 79, 3116-3129 11 Falini,B., Bolognesi, A., Flenghi, L. et al. (1992) Lancet339, 1195-1196 12 LeMaistre, CF., Meneghetti, C., Rosenblum, M. etal. (1992) Blood 79, 2547-2554
13 LeMaistre, C.F., Rosenhlum, M.G., Reuben, J.M. et a/. ( 1991 ) l,ancet 337, 1124-1125 14 Rybak, S.M., Saxena, S.K., Ackerman, E.J. and Youle, R.J. ( 199 I I. Biol. Chem. 266, 21202-21207 15 Wiley,R.G., Oeltma/m, T.N. and Lappi, D.A. ( 1991 / Brain Res. 562, 149-153
v,,t 1~ ,~,, to 1.,,,_,
Next month in IT... * Overview:
intrathymic and e x t r a t h y m i c T-cell
development * Candidate NK cell receptors * Evolution of
germinal centres * CD40 and its ligand plus...
* Centrepage diagram: cytokines in parasitic infections
F.W, (1989) I. Immunol. 143, 15-22 12 Coffman, R.L., Chatelain, R., Leal, L.M.C.C. and Varkila, K. (1991) Res. Immunol. 142, 35-40 13 LeGros, G., Ben-Sasson, S.Z., Seder, R., Finkelman, F.D. and Paul, W.E. (1990) I. Exp. Med. 172, 921-930 14 Locksley, R.M., Heinzel, F.P., Holaday, B.G. et al. (19911 Res. ImmunoL 142, 28-31 15 Maggi, E., Parronchi, P., Manetti, R. et al. (1992) J. Immunol. 148, 2142-2147 16 DeMaeyer, E. and DeMaeyerGuignard, J. (1988) Interferons and Other Regulatory Cytokines, John Wiley & Sons 17 Parronchi, P., De Carli, M., Manetti, R. et al. J. Immunol. (in press)
18 Kobayashi, M., Fitz, L., Ryan, M. et al. (1989)J. Exp. Med. 170, 827-845 19 Finkelman, F.D., Svetic, A., Gresser, I. et al. (1991)I. Exp. Med. 174, 1179-1198 20 Paul, W.E. (1992) in New Advances on Cytokines (Romagnani, 5., Abbas, A.K. and Mossman, T.R., eds), pp. 151-154, Raven Press 21 Ben-Sasson, S.Z., LeGros, G,, Conrad, D.H., Finkelman, F.D. and Paul, W.E. (1990)Proc. Natl Acad. Sci. USA 87, 1421-1425 22 Piccinni, M-P., Macchia, D., Parronchi, P. et al. (1991) Proc. Natl Acad. Sci. USA 88, 8656-8660 23 Conrad, D.H., Ben Sasson, S.Z., LeGros, G., Finkelman, F.D. and Paul W.E. (1990) 1. Exp. Med. 171, 1497-1508
Immunotoxins: the power and the glory Edward J. Wawrzynczak and Elaine J. Derbyshire
The traditional triumvirate of natural toxins - diphtheria toxin (DT), Pseudomonas exotoxin A (PE) and ricin - all act by binding to cells, undergoing internalization and intracellular processing; this allows ddivery to the cytoso[ of a catalytic component capable of inactivating protein synthesis. The secret of making a useful immunotoxin is to eliminate the cellbinding ability of the toxin without loss of the other functions vital to its action. Determination of the crystal structure of DT, presented by S. Choe (National Institutes of Health, Bethesda) ~, revealed a Y-shaped molecule with clearly delineated domains involved in receptor binding, translocation and catalysis. The receptor-binding domain is a flattened 13-barrel reminiscent of the immunoglobulin (Ig) fold. It was suggested that complete replacement of the receptor-binding domain by an Ig variable domain (F0 could redirect the binding specificity of DT.
Immunotoxins are hybrid proteins in which the potent cytocidal action of a toxin is harnessed for the selective destruction of target cells by attachment to a specific monoclonal antibody (mAb) or growth factor. This brief article describes the latest advances in the molecular and cellular biology, pharmacology and clinical evaluation of immunotoxins, as discussed at a recent meeting.
Elucidation of the DT structure has helped to rationalize experimental findings with recombinant single-chain hybrid fusion proteins between DT analogues and growth factors produced in Escherichia coll. L-F. Jean, F. Lakkis (University Hospital, Boston) and J. Shaw (Seragen Inc., Hopkinton) reported The Third International Symposium on lmmunotoxins was organized by Arthur E. Frankel and was held at the Sheraton World Resort, Orlando, Florida, USA on 19-21 June 1992.
the properties of fusions between interleukin 4 (IL-4), 1L-6 and epidermal growth factor (EGF) with DAB389, a truncated DT analogue. DAB3s9 completely lacks the recepto>binding domain and has superior targeting properties compared with DAB4s6, from which only part of the receptor-binding domain has been excised 2. The complexity of the mechanism of action of PE has been revealed in a series of elegant experiments by D. FitzGerald, I. Pastan and colleagues (National Institutes of Health, Bethesda). PE binds to the ubiquitous c~macroglobulin receptor. Following endocytosis, the toxin is cleaved after Arg279 by a cellular protease, to generate a cell-binding fragment and a catalytic fragment which becomes translocated to the cytosoP. Trp281 appears to be essential to an interaction with an intracellular component after the cleavage step. Gly280, the amino terminus of the catalytic fragment,
1992, Elsevier Science Publishers lid, UK.
Immunology Today
381
vo~. ~3 No. 1019v2
TRENDS
can be changed to Met280 without influencing translocation, allowing the creation of novel PE analogues no longer requiring intracellular proteolysis, a potential variable in the sensitivity of cells to immunotoxin. C. Siegall (Bristol-Myers Squibb, Wallingford) and U. Brinkmann (National Institutes of Health, Bethesda) described preclinical studies of recombinant single chain Fv fusions with truncated forms of PE lacking the cell binding domain, BR96(single-chain Fv)-PE40 and B3(single-chain Fv)-PE38KDEL. Both immunotoxins induced complete remission of large solid subcutaneous tumour xenografts in nude mice. The recombinant immunotoxins were superior in vivo to immunotoxin analogues made by chemical conjugation because of higher potency, lower toxicity, and better turnout penetration4. The attachment of high-affinity ligands to the two galactosebinding sites of ricin results in a blocked ricin with 1 000-fold lower cell binding, but there is evidence that the residual binding capacity is essential for the activity of blocked ricin immunotoxinss. R. Youle (National Institutes of Health, Bethesda) reported collaborative studies with M. Lord (University of Warwick) using recombinant ricin molecules in which either one or both binding sites had been incapacitated by site-directed mutagenesis. Wild-type and mutant recombinant ricin B (binding) chains were expressed in Xenopus oocytes and reassociated with recombinant ricin A (active) chain. The mannosyl side chains of the B chain mediated uptake of the reconstituted toxins by macrophages. Removal of both binding sites abolished toxicity, indicating the importance of galactose-binding in efficient delivery of the A chain to the cytosol6. The A chain of ricin can itself be targeted by direct attachment to mAb via a disulphide bond. A variety of other ribosome-inactivating proteins (RIPs) from plants and fungi, including gelonin, saporin, trichosanthin and mitogillin, have been cloned and expressed in E. coli. S. Bernhard (XOMA Corp.,
B-cell lymphoma using an antiCD19 immunotoxin, HD37-deglycosylated ricin A. In a severe combined immunodeficiency (SCID) mouse model of disseminated Bcell lymphoma developed by M-A. Ghetie (University of Texas, Dallas) 9, a mixture of the antiCD22 immunotoxin and either the anti-CD19 immunotoxin or the CD19 mAb gave superior antitumour effects, providing a rationale for combination therapy. Clinical trials Immunotoxins made with natuH65-ricin A chain (now known under the trade name 'CD5 Plus'), ral RIPs have recently reached the an immunotoxin that was directed stage of clinical evaluation. B43antiviral against the CD5 antigen, has (anti-CD 19)-pokeweed been administered to hundreds of protein, developed by F. Uckun patients with graft-versus-host dis- and colleagues (University of ease (GVHD) following allogeneic Minnesota, Minneapolis), demonbone marrow transplantation. P. strated significant anti-tumour Scannon (XOMA Corp., Berkeley) effects in a SCID mouse model, reported that when two groups especially in combination with cyof patients with steroid-resistant clophosphamide ~°, and has recently GVHD were treated with the entered phase I trials. B. Falini immunotoxin, enhanced resol- (University of Perugia, Italy) reution of acute GVHD occurred and ported early results of trials in there was a survival benefit in patients with refractory Hodgkin's grade I-III GVHD compared with disease treated with an anti-CD30 historical controls. Studies are in immunotoxin, Ber-H2-saporin J~. progress with 'CD5 Plus' in F. LeMaistre (University of patients with rheumatoid arthritis, Texas, San Antonio) and T. type I diabetes mellitus and other Woodworth (Seragen Inc., Hopkinautoimmune diseases. ton) reported trials with the first M. Grossbard (Dana-Farber recombinant immunotoxin in the Cancer Institute, Boston) pre- clinic, DAB486IL-2. This immunosented the results of a number of toxin selectively eliminates tumour trials using anti-B4-blocked ricin, cells and activated T cells that directed against the CD19 pan-B- express the high affinity IL-2 cell antigen, in patients with B-cell receptor and is thus applicable to a neoplasms refractory to conven- variety of haematological maligtional treatments7. A phase I trial nancies 12,~3, rheumatoid arthritis of adjuvant therapy for patients and insulin-dependent diabetes with non-Hodgkin's lymphoma mellitus. Future trials are planned following autologous bone marrow with another recombinant immunotransplantation is in progress. toxin, DAB3s9IL-2. E. Vitetta (University of Texas, Although many trials are still at Dallas) presented the results of a a preliminary stage, it is possible to collaborative study among several draw some general conclusions. centres comparing the behaviour of First, all immunotoxins show a two deglycosylated ricin A chain similar (though not identical) patimmunotoxins linked either to the tern of transient and reversible anti-CD22 mAb RFB4 or its Fab' toxicities; primarily, an elevation fragment, in phase I trials in of liver transaminases and/or a patients with B-cell lymphoma8. vascular leak syndrome characterDespite differences in potency, ized by hypoalbuminaemia and pharmacokinetics, toxicity and im- oedema. Secondly, a number of munogenicity, the overall perform- long-lasting remissions were ance of both agents was similar. achieved in dose-escalation studies M. Stone (Baylor-Sammons Cancer and anti-tumour effects were seen Center, Dallas) reported prelimi- in heavily pretreated patients. nary data of a new phase I trial in Thirdly, host antibody responses to Berkeley) used mutagenesis of recombinant single chain RIPs to create analogues with a single, free thiol group for site-specific conjugation to mAb. The position of the thiol on the RIP influenced enzymic activity, conjugation efficiency and immunotoxin potency; some RIP analogues formed more potent immunotoxins than the natural RIP or ricin A chain.
Immunology Today
382
Vol 13 No. 10 •992
TRENDS
immunotoxins occurred in many patients, even in a proportion of immunosuppressed lymphoma patients.
mAbs that recognize human antigens associated with tumour neovasculature, which could be applied to many different types of tumour. Although much of the focus of Novel approaches immunotoxin research is directed The immune response to murine towards alleviation of human lg and toxin molecules of bacterial disease, immunotoxins are also or plant origin limits the amount of proving to be useful experimental immunotoxin that can be admin- tools. R. Wile}' (DVAMC and istered to patients. D. Newton Vanderbilt University, Nashville) (National Institutes of Health, described the use of immunotoxins Bethesda) found that bovine pan- to ablate specific neurones in excreatic RNAse linked to an anti- perimental animals. A saporin transferrin receptor mAb displayed immunotoxin directed against the selective cytotoxic activity against rat low-affinity neurotrophin receptumour cells in tissue culture L4. tor ~s was highly effective in Despite considerably lower potency destroying peripheral neurones than an analogous ricin A chain when administered systemically to rats. Microimmunotoxin, the RNAse immuno- Sprague-Dawley toxin exerted similar growth in- injection into the anterior lateral hibitory effects when injected into ventricle selectively destroyed neuglioblastoma xenografts. RNAses rones of the cholinergic basal foreof mammalian or human origin are brain. Since these are the neurones likely to be much less immunogenic that predominantly degenerate in than the conventional toxins, es- Alzheimer's disease, this rat model pecially in combination with may be helpful in investigating the pathophysiology of the disease and human or humanized mAbs. Conventional immunotoxins in testing potential therapies. readily reach turnout cells close to blood capillaries but penetrate Conclusion The rapid advances in our undersolid tumour masses poorly. A complementary approach to killing standing of toxin structure and malignant cells directly is to target action, new techniques for molecuthe vasculature upon which the lar engineering of immunotoxins, tumour relies for its nutrient promising results of recent patient supply. P. Thorpe and F. Burrows studies, novel approaches, and (University of Texas, Dallas) expanding interest suggest that a presented a mouse model of vascu- glorious future lies ahead for the new generation of lar targeting. The neuroblastoma powerful cell line C1300, transfected with immunotoxins. the gamma-interferon (IFN-y) gene, was grown in BALB/c nude mice. Eddie Wawrzynczak is at the IFN-y secreted by the tumour Institute o f Cancer Research, induced the expression of major Sutton, Surrey, UK SM2 5NG; histocompatibility complex (MHC) Elaine Derbyshire is at the class II antigens on the endothelial University o f Texas Southwestern cells adjacent to the turnout. An Medical Center, Dallas, T X 75235anti-class I1 immunotoxin elicited 8576, USA. extensive turnout necrosis through a process of endothelial cell damage, platelet deposition and References intravascular thrombosis; only a 1 Choe, S., Bennett, M.J,, Fujii, G. thin rind of turnout cells adjacent et aL (1992) Nature 357, 216-222 to normal skin blood vessels 2 Shaw, J.P., Akiyoshi, D.E., Arrigo, survived the immunotoxin treat- D.A. et al. (1991).I. Biol. Chem. 266, ment. When the treatment included 21118-21124 3 Fryling, C., Ogata, M. and a second immunotoxin directed FitzGerald, D. (1992) Infect. lmmun. against the tumour itself, the 60,497-502 readily accessible cells were also 4 Brinkmann, U., Pai, L.H., killed and complete turnout re- FitzGerald, D.J. et al. (1991) Proc. Natl gressions resulted. The hunt is on for Acad. Sci. USA 88, 8616-8620
t,,,,,,,ology r,,aa,,
383
5 Goldmacher, V.S., Lambert, J.M. and Blattler, W.A. (I 992) Biochem. Biophys. Res. Commun. 183~ 758-766 6 Newton, D.I,., Wales, R., Richardson, P.T. et al. (1992)1. Biol. Chem. 267, 11917-11922 7 Grossbard, M.L., Freedman, A.S., Ritz, J. et al. (1992) Blood 79, 576-585, 8 Vitetta, E.S., Stone, M., Amlot, P. et al. (1991) Cancer Res. 5 I, 4052-4058 9 Ghetie, M-A., Richardson, J., Tucker, T. et al. (1991) Cancer Res. 51, 5876-5880
10 Uckun, F.M., Chelstrom, L,M., Finnegan, D. et al. (1992) Blood 79, 3116-3129 11 Falini,B., Bolognesi, A., Flenghi, L. et al. (1992) Lancet339, 1195-1196 12 LeMaistre, CF., Meneghetti, C., Rosenblum, M. etal. (1992) Blood 79, 2547-2554
13 LeMaistre, C.F., Rosenhlum, M.G., Reuben, J.M. et a/. ( 1991 ) l,ancet 337, 1124-1125 14 Rybak, S.M., Saxena, S.K., Ackerman, E.J. and Youle, R.J. ( 199 I I. Biol. Chem. 266, 21202-21207 15 Wiley,R.G., Oeltma/m, T.N. and Lappi, D.A. ( 1991 / Brain Res. 562, 149-153
v,,t 1~ ,~,, to 1.,,,_,
Next month in IT... * Overview:
intrathymic and e x t r a t h y m i c T-cell
development * Candidate NK cell receptors * Evolution of
germinal centres * CD40 and its ligand plus...
* Centrepage diagram: cytokines in parasitic infections
