Immunology 1988 65 329-335
`
`REVIEW PAPER
`
`Therapeutic strategies with monoclonal antibodies and
`immunoconjugates
`
`V. S. BYERS*t & R. W. BALDWIN* * Cancer Research Campaign Laboratories, University of Nottingham, Nottingham,
`U.K. and t Xoma Corporation, Berkeley, California, U.S.A.
`
`Acceptedfor publication I August 1988
`
`CONTENTS
`
`Therapeutic use of mAbs
`Immunotoxins
`Chemo-immunoconjugates
`Clinical trials
`Human anti-murine monoclonal antibody responses (HAMA)
`Clinical impact
`Approaches to abrogate anti-mouse immunoglobulin responses in patients
`Conclusions
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`Introduction
`Monoclonal antibodies (mAbs) are emerging as a major
`modality for both detection and therapy ofvarious pathological
`conditions, including malignant disease. Murine monoclonal
`antibodies have been developed which react with tumour-
`associated antigens expressed on or near the cell surface, and
`present in high concentration on malignant tissues but in
`minimal amounts on normal tissues (Boyer et al., 1988). These
`include mAbs against colon cancer, ovarian cancer, breast
`cancer and neurological tumours, which have been used thera-
`peutically either alone or conjugated to cytotoxic agents
`(Baldwin & Byers, 1987) and diagnostically coupled to radio-
`isotopes for gamma camera imaging of patients (Chatal, 1988,
`Pimm, 1987). More recently investigators have taken advantage
`ofthe well-defined antigens on the surface oflymphoid cells, and
`mAbs directed against such antigens have been used therapeuti-
`cally against lymphoid malignancies and autoimmune diseases,
`including prevention of renal allograft rejection and treatment
`of graft-versus-host (GVH) disease (Goldstein et al., 1985;
`Byers, 1987; Byers et al., 1987a; Fahey et al., 1987; Youle &
`Colombatti, 1986).
`In spite of the encouraging clinical results, including the use
`of the anti-T-cell receptor mAb OKT3 in renal allograft
`rejection (Goldstein et al., 1985; Mayes et al., 1988), the central
`limitation in using these murine products has been the gene-
`ration of an immune response to murine immunoglobulin which
`usually precludes retreatment. Especially in solid tumour
`therapy, retreatment is viewed as a key factor in allowing this
`therapeutic modality to be optimized, and a variety of
`approaches to solve the problem are underway.
`These issues are reviewed in relation to the now quite
`extensive advances in designing immunoconjugates for therapy,
`particularly in malignant disease.
`Correspondence: Dr V. S. Byers, Cancer Research Campaign
`Laboratories, University of Nottingham, Nottingham NG7 2RD, U.K.
`
`Therapeutic use of mAbs
`Murine monoclonal antibodies that recognize tumour-asso-
`ciated antigens and localize in tumours, or that have well-
`defined reactivity against antigens on lymphocytes, are being
`evaluated for therapy (Baldwin & Byers, 1987). Unmodified
`antibodies have demonstrated efficacy in some cases. For
`example, in renal allograft transplantation OKT3 antibody
`directed against the T-cell receptor has been proven quite
`effective in reversing rejection, probably as a result of blocking
`lymphocyte-antigen interactions (Goldstein et al., 1985). Also,
`unconjugated antibodies are being used in cancers such as colon
`cancer (LoBuglio et al., 1986; Sears et al., 1985) and neuroblas-
`toma (Cheung et al., 1987). The rationale for their use is that
`host defence mechanisms such as antibody-dependent cell
`cytotoxicity (ADCC) can serve as the cytotoxic modality, but
`consideration is now being given to the view that murine
`monoclonal antibodies may be inducing anti-idiotypic anti-
`bodies that will react against the tumour (Viale et al., 1987;
`Nepom & Hellstrom, 1987; Herlyn et al., 1987a, b). These
`approaches have shown some encouraging results-for example
`in the case of antibodies against neuroblastoma-but most
`attention has turned to the use of mAbs for targeted therapy
`using cytotoxic agents, including radioisotopes and toxins.
`The initial selection ofantibody for use as a targeting agent is
`primarily based upon its reactivity with appropriate target cells
`in comparison with binding to normal cells, since this latter
`reactivity will be translated in vivo into 'side-effects'. This is still
`a major limitation, although there are only a few antibodies
`which meet specificity requirements. For example, with solid
`tumours, preclinical studies indicate that antibodies which react
`with malignant melanoma (Spitler et al., 1987) and colorectal
`cancer (Baldwin & Byers, 1988; Byers et al., 1988a, b) had
`minimal normal tissue reactivity, and immunotoxins made from
`these monoclonal antibodies were well tolerated in clinical
`studies. However, other potentially useful antibodies, such as
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`V. S. Byers & R. W. Baldwin
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`those binding to epidermal growth factor receptor (Griffin et al.,
`1987; FitzGerald et al., 1987) and to breast/ovarian cancer-
`associated mucins (Price, 1988a, b), exhibit quite widespread
`normal tissue reactivity. Immunotoxins constructed with these
`antibodies may exhibit normal tissue toxicity, thus limiting their
`clinical use.
`The mechanism of action of the different cytotoxic agents
`influences to some extent the choice of antibody. With radio-
`isotopes, the monoclonal antibody must be targeted to the
`tumour, but internalization into the cell may be detrimental
`since it can result in deconjugation of the isotopes from the
`monoclonal antibody, and exocytosis, facilitating its removal
`from the tumour (Press et al., 1988). On the other hand, most
`cytotoxic drugs, e.g. methotrexate, are bound to the antibody by
`labile bonds and endocytosis into an acidic intracellular com-
`partment maximizes activity through drug release (Garnett et
`al., 1985; Embleton & Garnett, 1986). Antibody targeting with
`these agents is intended primarily to reduce uptake by normal
`tissues, so lowering toxicity, although it may convert resistant
`cells to susceptible cells in cases where resistance is due to
`defective drug uptake. In contrast, ricin toxin A chain (RTA) is
`inactive extracellularly and non-toxic as the free agent. Anti-
`body conjugation targets the toxin moiety to the target cell,
`allowing it to be internalized by endocytosis (Baldwin et al.,
`1988; Byers et al., 1988a).
`
`Immunotoxins
`Toxins of plant and bacterial origin represent a class of highly
`cytotoxic agents for linking to monoclonal antibodies to form
`immunotoxins (Vitetta et al., 1987; Baldwin & Byers, 1987;
`Blakey & Thorpe, 1987). One type of toxin, typified by ricin
`produced from beans of the plant Ricinus Communis, is
`composed of two disulphide-linked subunits. The intact toxin
`binds to mammalian cells through galactose residues on the B
`subunit and is endocytosed (Van Deurs et al., 1988). The A
`chain, which is inactive extracellularly, after introduction into
`the cell catalytically inactivates the 60s ribosomal subunit of
`eukaryotic cells by modifying one or two nucleoside residues of
`the 28s ribosomal RNA (Endo et al., 1987). If the bond between
`the A and B chains is broken and an antibody substituted for the
`B chain, the A chain can now be directed to specific target cells.
`This approach has now been used for several A-B chain toxins,
`including pseudomonas exotoxin, diphtheria toxin and abrin
`(Vitetta et al., 1987; Baldwin & Byers, 1987).
`Much work has been done with RTA immunotoxins
`containing ricin A chain conjugated to antibody through
`the cross-linker N-succinimidyl-3-(2-pyridyldithio) propionate
`(SPDP). This contains a disulphide bond, and it is generally
`assumed that this is cleaved intracellularly to allow the A chain
`to translocate through the cytoplasm to the Golgi apparatus.
`The pharmacokinetics of the clearance of the antibody moiety
`of the immunotoxins is markedly changed by the addition of the
`RTA (Byers et al., 1987b; Bourrie et al., 1986; Blakey et al.,
`1987) since this protein contains mannose-terminating oligosac-
`charides that are recognized by reticuloendothelial cells, includ-
`ing Kupffer cells in the liver (Blakey et al., 1987). This results in
`rapid liver clearance and reduces the blood half-life from days to
`hours. Increasing the blood half-life improves localization of
`immunotoxin in the tumours, so various strategies are being
`devised to eliminate the mannose residues on the RTA without
`affecting activity. These include deglycosylation of RTA
`
`(Blakey et al., 1988), synthesis of recombinant RTA, and use of
`a subfraction of RTA with reduced oligosaccharide content
`(Fulton et al., 1986). The increased circulation time increases
`localization at the disease site; this must be balanced against the
`possibility of increased or altered toxicity.
`Endocytosis and intracellular processing are essential for
`immunotoxins to exert their effect (Vitetta et al., 1987; Baldwin
`et al., 1988). Since the mechanism of intracellular trafficking is
`poorly understood, immunotoxins are selected by screening
`them for specific cytotoxicity. This is usually expressed as the
`concentration of immunotoxin necessary to inhibit 50% of
`target cell growth in vitro (IC50) and, in most cases, the difference
`in potency between immunotoxin and free RTA is in the order
`of 103-104-fold. One immunotoxin against the CD22 antigen on
`B lymphomas actually has a lower ICo than whole ricin (May et
`al., 1986).
`As additional information of the mechanism of internaliza-
`tion and intracellular trafficking of immunotoxins is obtained,
`construction of potent immunotoxins should become more
`efficient. Both the rate of endocytosis and the intracellular
`compartment into which the immunotoxin is moved depends in
`some part on the antigen to which the monoclonal antibody
`binds. Receptor-mediated endocytosis is more efficient than
`pinocytosis, for example the rate of endocytosis of a gelonin-
`containing immunotoxin which reacts with the C3b receptor on
`human B lymphocytes can be markedly increased by infection
`with Epstein-Barr virus, which is the preferred ligand for the
`receptor (Tedder et al., 1986). Also, recent studies by Sandvig et
`al. (1987) demonstrated that under conditions in which recep-
`tor-mediated endocytosis through coated pits is blocked, inter-
`nalization of transferrin and epidermal growth factor does not
`occur, but internalization and intoxication by ricin continues
`essentially unhindered. These and other data have been taken to
`suggest that at least two independent mechanisms exist for the
`internalization of surface-bound ligands, one via coated pits,
`and the other via 'smooth' pits which lack a clathrin coat. This
`may influence the cellular compartment into which the immuno-
`toxin is directed. After endocytosis, the endocytotic vacuole is
`acidified and, in contrast to chemoimmunoconjugates and
`immunotoxins such as diphtheria toxin, RTA immunotoxins
`are inactivated by acidic environments. Thus for these conju-
`gates, any inhibition of intracellular acidification increases
`activity. The potent potentiating agents for many immunotox-
`ins, monensin and ammonium chloride, are postulated to work
`by decreasing the rate of intracellular acidification, thereby
`prolonging intracellular activity of the immunotoxins.
`
`Chemo-immunoconjugates
`The design of chemo-immunoconjugates is developing along
`similar lines to those established with immunotoxins, and
`conjugates have been produced with several classes ofcytotoxic
`drugs, including alkylating agents and antimetabolites (Bald-
`win, 1985; Baldwin et al., 1986; Ghose & Blair, 1987). These are
`produced by conjugating drug to antibody through a specific
`functional group such as amino, hydroxyl or sulphydryl
`residues. In this case, the selected conjugation site should not be
`required for drug action or, if so, should become available
`following intracellular release of the drug. Cytotoxic drugs are
`generally less active than toxins and it is necessary to introduce
`the maximum number of drug residues compatible with reten-
`tion of antibody reactivity. High drug substitution ratios have
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`been reported, but with monoclonal antibodies of the IgG
`subclass, substitution of more than 10 drug residues per
`antibody molecule may produce an unacceptable level of
`antibody damage. Consequently drug carrier molecules such as
`human serum albumin and dextrans are being used to yield
`antibody conjugates with much higher molar ratios of drug to
`antibody (Garnett & Baldwin, 1986; Endo et al.,
`1988).
`Conjugates of large molecular size may have inappropriate
`pharmacokinetic properties and are less able to penetrate
`tumour tissue. Currently, therefore, attention is being given to
`the chemistry of drug-antibody conjugation so as to improve
`the potency of chemo-immunoconjugates. This includes the
`selection of more cytotoxic analogues of the drugs used for
`antibody conjugation, and design of new drug carrier molecules
`to replace currently used polymers, as well as modification ofthe
`drug conjugates, particularly the overall charge in relation to
`their pharmacokinetics.
`The use of hybrid antibodies is being considered as an
`alternative to chemical conjugation for in vivo targeting drugs to
`tumours. Hybrid-hybrid monoclonal antibodies recognizing
`both a tumour-associated antigen and the drugs are being
`produced either by fusion of two existing hybridomas or by
`fusion of one existing hybridoma with spleen cells from donor
`mice immunized against the other antigen. Studies in this field
`have been reported by Corvalan et al. (1987a, b) in which a
`hybrid antibody reactive with a carcinoembryonic antigen
`(CEA) epitope and vinca alkaloid was produced by fusion of an
`anti-CEA antibody-producing hybridoma with spleen cells
`from mice immunized with vindesine-protein conjugate. The
`IgG antibody had the yl heavy chain from the parental anti-
`CEA antibody and a y2a from the anti-vinca alkaloid donor
`lymphocyte. This antibody localized in an appropriate CEA
`producing colon carcinoma xenograft. The antibody also
`induced profound changes in the biodistribution of 3H-labelled
`vinblastine, resulting in specific localization of the drug to
`tumour tissue and so producing a therapeutic response.
`It is feasible to prepare monoclonal antibodies to other
`drugs and therefore there is the possibility of constructing a
`range of hybrid antibodies. For example, Pimm et al., (1987)
`with a hybridoma producing monoclonal antibody to metho-
`trexate showed that complexing antibody with drug profoundly
`altered the biodistribution of the drug, prolonging blood
`survival and reducing liver uptake, and clearly hybridomas such
`as this are candidates for constructing hybrid-hybrid antibodies
`with anti-tumour antibodies.
`
`Clinical trials
`Clinical trials are in progress with murine monoclonal anti-
`bodies alone and as immunoconjugates. Antibodies specific for
`the idiotype of the surface immunoglobulin of B-cell lympho-
`mas have been used successfully in inducing tumour regression,
`but the clinical outcome has been widely variable ranging from
`prolonged remission to no effect (Lowder et al., 1987). Factors
`involved include antigen modulation, which may be of particu-
`lar concern with lymphoid tumours, and generation of anti-
`mouse antibody responses which markedly alter antibody
`pharmacokinetics. Another approach with B-cell tumours
`involves treatment with murine monoclonal antibodies directed
`against normal CD20 antigens. Treatment of four patients with
`refractory B-cell lymphomas with an anti-CD20 antibody
`
`produced a 90% elimination ofmalignant cells from the blood in
`two patients and a reduction in lymph node disease in one
`patient, but the duration of remission was shortlasting (Press et
`al., 1987). Antibodies specific for tumour-associated ganglio-
`sides which activate human complement and are active in
`ADCC are in clinical trials with neuroblastoma (Cheung et al.,
`1987). The antigen recognized by this antibody is also expressed
`on neurons and peripheral brain fibres. Because of pain during
`infusion, patients must be anaesthetized and maintained on
`analgesics for several weeks afterwards. However, complete
`remission of two neuroblastomas have been reported, suggest-
`ing that, with appropriate medical support, substantial reacti-
`vity with normal tissues can be tolerated.
`The most successful therapeutic use of an unconjugated
`antibody has been the use of OKT3 in renal allograft rejection
`(Goldstein et al., 1985). This murine monoclonal antibody is
`directed against the CD3 (T-cell receptor) antigen on mature
`human T cells and blocks their function. When given over a 14-
`day period as initial therapy in patients experiencing acute
`rejection, it will reverse 94% of the rejections, and significantly
`improve I year graft survival to 62%. This is now an approved
`drug for that indication.
`Clinical evaluation of radioisotope-labelled monoclonal
`antibodies is in progress, using 131-iodine (gamma and B
`emission), 90-yttrium (gamma emission) and 21 1-astadine (a-
`emission) (Humm, 1986; Order et al., 1988; Sands, 1988). It has
`been argued that the minimum requirement for effective therapy
`using radioimmunoconjugates is accumulation of 10 times more
`conjugate in tumour compared with blood (Vaughan et al.,
`1987) and this cannot be attained by intravenous administra-
`tion. Therefore, recent clinical applications have focused upon
`intraperitoneal injection of 131 -iodine-labelled antibody in
`patients with ovarian and colon cancer and intrathecal injection
`in patients with malignant melanoma. In one trial in ovarian
`cancer (Stewart et al., 1988) patients were treated with a range of
`antibodies. There were complete responses in 3/6 patients with
`microscopic disease and partial response in 2/15 patients with
`nodules smaller than 2 cm.
`The most impressive response to immunotoxin therapy so
`far has been in the treatment of acute graft versus host (GVH)
`disease developing following bone marrow transplantation
`(Kernan et al., 1988; Byers et al., 1987b). The immunotoxin used
`contains RTA conjugated to a monoclonal antibody reacting
`with the CD5 antigen on mature T lymphocytes. In 12/25
`evaluable patients, progression of disease was reversed after the
`first seven doses of immunotoxin; in another five it was
`stabilized. Overall these patients continued to attain complete
`responses as late as 60 days after therapy, whereas most of the
`non-responders had died by Day 40. Responses were seen in all
`three involved organs, skin, gut and liver. Pharmacokinetic
`studies indicated that the immunotoxin was cleared rapidly
`from blood. Even so, the circulating T lymphocytes, as mea-
`sured by CD3 and CD5 expression, dropped by Day 6 of
`treatment to less than 20% of the initial value and remained low
`for several weeks to months thereafter.
`Clinical trials with ricin A chain immunotoxins also include
`phase I/II trials in malignant melanoma with antibody directed
`against the high molecular weight (> 200,000) melanoma
`antigen (Spitler et al., 1987) and a phase I trial in colorectal
`carcinoma with monoclonal antibody 791T/36 which reacts
`with a 72,000 molecular weight tumour membrane glycoprotein
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`(gp72) (Byers et al., 1988b). These immunotoxins are well
`tolerated in terms of side-effects. These include reversible
`hypoalbuminemia, and myalgias, which are generic for the
`immunotoxins, and proteinuria which is associated with the
`immunotoxin constructed with monoclonal antibody 791T/36.
`In the melanoma study, approximately 50% of the patients
`showed some degree of response, with one patient having
`complete remission. In the colorectal cancer study, three
`patients with hepatic metastases had resolution of the smaller
`lesions. Although encouraging, most of the clinical responses
`have not yet reached the requirement for categorization as
`response by the WHO criteria. For this to be achieved it is likely
`that retreatment schedules will be necessary and for this the
`development of anti-mouse antibody responses in immunotoxin
`patients must be controlled.
`
`Human anti-murine monoclonal antibody responses (HAMA)
`Clinical impact
`Almost all of the murine monoclonal antibodies being used
`clinically provoke antibody responses (HAMA) in patients.
`These include antibody responses against the framework, the
`isotype, and the idiotype of the murine IgG antibodies. It is rare
`that anaphylaxis or serum sickness occurs in patients with
`HAMA when retreated with mAb, although anaphylactoid
`reactions are not uncommon and primarily consist ofperiorbital
`oedema or urticaria. But HAMA responses lead to altered
`pharmacokinetics of the injected monoclonal antibody. Anti-
`melanoma immunotoxin is rapidly cleared from serum with
`reduced drug levels being attained (LoBuglio et al., 1988a).
`Immunoscintigraphy of tumours with radiolabelled antibody is
`also seriously compromised in patients developing HAMA, and
`has resulted in negative imaging of tumour with radiolabelled
`product (immune complexes) being localized in liver and spleen
`(Pimm et al., 1985; Perkins, Pimm & Powell, 1988). The extent to
`which HAMA influences therapy clinically is still unclear. One
`study of patients treated with OKT3 (Jaffres et al., 1986)
`reported that of 21 patients given OKT3 60% made anti-Id
`antibodies, which in some cases interfered with its therapeutic
`effectiveness, while another report indicated that low levels of
`blocking antibodies did not significantly compromise access to
`OKT3 for treatment of subsequent rejection episodes (Mayes et
`al., 1988). Possibly the more important finding is the fact that
`even though these patients were immunosuppressed with azath-
`ioprine, prednisone, and in some cases cyclosporine, they still
`were able to generate HAMA with anti-Ids as a prominent
`feature of the response, even though this must constitute a
`relatively minor portion of the murine monoclonal antibody.
`The actual generation of HAMA will probably depend in some
`part on the underlying disease as well as the concomitant
`immunosuppression; for example patients with cutaneous T-cell
`lymphoma have a higher incidence of HAMA to the anti-CD5
`mAb T1O1 than do patients with CLL, where almost no patients
`produce an immune response (Dillman et al., 1984). Also,
`patients receiving an immunotoxin composed of an anti-CD5
`mAb coupled to RTA, who are suffering from GVH disease and
`therefore are immunosuppressed both exogenously and endoge-
`nously, have very little HAMA (V. S. Byers, R. P. Mischak, N.
`Kernan and P. Scannon, unpublished findings). In some
`situations, a single course of mAb therapy may be sufficient to
`cure the disease, analagous to the experience with anti-thymo-
`
`cyte globulin in which 4-10 days of therapy is sufficent to cure
`aplastic anaemia (Young et al., 1988). Overall, however, most
`investigators take the position that if multiple courses of mAb
`are to be used therapeutically, reliable methods for abrogating
`the immune response must be devised.
`
`Approaches to abrogate anti-mouse immunoglobulin responses in
`patients
`Concomitant treatment with immunosuppressive medication is
`one method being developed, and in one study the primary
`HAMA response was blunted following pretreatment with
`azathioprine/prednisone for 8 weeks (LoBuglio et al., 1988). A
`significant inhibition of the primary HAMA response has been
`achieved with cyclosporin (CsA) pretreatment for 6 days,
`although CsA had no effect on abrogation of response in
`patients with pre-existing anti-murine antibodies (Lederman et
`al., 1988). The problems with such regimens is non-specific
`immunosuppression, and toxicity of the additional drugs which
`makes such therapy with mAbs more hazardous than with mAb
`given as a single agent. An alternative approach is to produce a
`human antibody or 'humanized' version of murine monoclonal
`antibodies.
`Initially, intensive efforts were directed toward making
`human mAbs, but products generated have been unsatisfactory
`(Campbell et al., 1987; James & Bell, 1987). Lymphocytes from
`human peripheral blood or from draining nodes from human
`tumours were fused with either human or murine myeloma
`partners, and the mAbs generated were of the IgM subclass and
`usually directed against intracellular antigens. IgM antibodies
`do not function well as imaging agents, probably because they
`are too large to marginate from the peripheral blood into the
`tissues, and would also be expected to be poor therapeutic
`agents for that same reason. The problem in finding mAbs
`directed against human cell membrane antigens may relate to
`humans being tolerant to such antigens, but the reason for the
`difficulty in producing human mAbs of the IgG subclass is still
`not understood. Meanwhile, however, methods are being
`devised by which the variable region of the murine mAb may be
`engineered onto a human constant region. A chimeric antibody
`composed of the variable regions of murine monoclonal
`antibody 17.1A (IgG2aK), which recognizes a glycoprotein
`present on human colorectal cells, has been coupled with the
`constant region of human IgG3. This chimeric 17.1A mAb has
`the same reactivity against colon tumour cells as the native
`antibody, and both have identical capacity to inhibit radiola-
`belled native 17. 1A binding to tumour cells (Shaw et al., 1987).
`Native 17.1A has been used extensively in clinical trials in
`colorectal cancer patients, and it elicits a very pronounced
`HAMA response, altering its pharmacokinetics (LoBuglio et
`al., 1988b; Khazael et al., 1988). In contrast, antibody responses
`have not been detected in patients receiving multiple treatments
`with the chimeric 17.1A, and the pharmacokinetics of the
`chimeric 17.1A was not modified, as additional evidence of lack
`of an antibody response (Shaw, Khazaeli & LoBuglio, 1988).
`This result was unexpected since this construct still has the
`whole of the variable region as murine derived, and since the
`idiotypic component of the immune response forms a prominent
`proportion of the HAMA response. Several groups are, there-
`fore, in the process of 'humanizing' murine mAbs, using various
`techniques such as the chimeric antibody constructed from rat
`antibody CAMPATH I by introducing only the six hypervari-
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`able regions from the heavy- and light-chain domains into a
`human IgGl molecule (Riechmann et al., 1988). This chimera
`was as effective as the native mAb in complement-mediated lysis
`of B-cell lymphocytic leukaemia cells.
`Control of the immune response to immunoconjugates will
`probably be more difficult, especially where the conjugated
`moiety itself is immunogenic, e.g. ricin A chain. The immuno-
`conjugates themselves should theoretically exert a cytotoxic
`effect not only against their target cell, but also against the
`antigen-specific cells of the immune system that are capable of
`producing antibody responses against them. Thus antigen-
`specific B lymphocytes having clonally distributed specific
`surface immunoglobulin receptors should react with immuno-
`conjugates. Such recognition should lead to endocytosis of the
`cytotoxic conjugate and, following intracellular release, the
`cytotoxic moiety should cause cell death. The HAMA seen with
`certain immunoconjugates, such as RTA-containing immuno-
`toxins, indicates that for various reasons they fail to adequately
`exert a cytotoxic effect against B lymphocytes. Immunoconju-
`gates have been constructed, however, which suppress anti-
`mouse antibody responses. Conjugates constructed with mono-
`clonal antibody 791T/36 and cis-aconityl-substituted daunomy-
`cin render rats immunologically unresponsive to subsequent
`repeated immunization with the unconjugated antibody (Dur-
`rant et al., 1988). However, rats still produced HAMA in
`response to challenge with the RTA immunotoxin made from
`791T/36 antibody. This probably means that RTA itself is
`immunogenic and serves as an effective antigen-presenting
`molecule, possibly through its avid uptake by macrophages.
`Supporting this view is the finding that murine monoclonal
`antibody 791 T/36 coupled to ricin A chain produces strong anti-
`idiotypic antibody responses in mice (Pimm, Durrant & Bald-
`win, 1988). Further manipulation may be necessary, therefore,
`to tolerize the host against the cytotoxic moieties such as ricin A
`chain prior to therapy with immunoconjugates constructed with
`humanized antibodies. Benjamin et al. (1988) showed that mice
`made tolerant to rat immunoglobulin constant region determi-
`nants were, in some instances, capable ofinducing only minimal
`reactivity to the idiotypic region of the antibody and the same
`approach may be used to suppress specific responses to
`immunoconjugates. If, however, there is a non-specific enhance-
`ment of antigen processing by components attached to huma-
`nized murine monoclonal antibodies, the pathway would be to
`generate cytotoxic analogues to eliminate this component. For
`the moment, therefore, investigators using immunoconjugates
`continue to utilize concomitant therapy with immunosuppres-
`sive agents such as CsA.
`
`Conclusions
`One should not underestimate the complexities of the problems
`still to be resolved in producing monoclonal antibodies and
`immunoconjugates for clinical use, especially in solid tumours.
`Outstanding problems include improving the design of the
`immunoconjugates themselves, particularly refining techniques
`for directing the conjugation site away from the active site of the
`antibody and conjugating adequate amounts of cytotoxic
`agents. More effective conjugation procedures are also impor-
`tant, particularly with small hydrophobic molecules. Second
`generation immunoconjugates, one anticipates, will be pro-
`duced with even better therapeutic efficacy.
`
`Enhanced target tissue accessibility is very important with
`solid tumours, since these are notorious for poor vascularity.
`Longer serum half-life will increase tumour localization, but it
`probably will be necessary to develop agents with a direct effect
`on tumour vascularity for adequate penetration. Antigenic
`heterogeneity is also an issue that is being addressed by use of
`cocktails of monoclonal antibodies, and it is still under discus-
`sion as to whether the cytotoxic molecules bound to the different
`monoclonal antibodies should also be different. More impor-
`tant is the issue ofantibody responses to the immunoconjugates.
`If these agents are viewed by the same criteria as conventional
`cytotoxic agents, it will be critical to retreat patients with more
`than one course of immunoconjugate, and possibly with
`immunoconjugates of different specificities. The antibody re-
`sponse seen with almost all monoclonal antibodies and immu-
`noconjugates at present effectively restricts treatment to a single
`short cycle. Several strategies have been identified to overcome
`this, including the design of human or humanized monoclonal
`antibodies. Whether or not this will restrict the generation of
`anti-idiotypic antibodies is not yet known, although studies with
`monoclonal antibody 17-1A suggest that in some cases this may
`be achieved. Even so it seems that immunoconjugates may still
`produce anti-idiotypic antibodies, and so in these cases immu-
`nosuppressive procedures may be needed. This may be achieved
`with immunosuppressive drug treatment, but with the asso-
`ciated toxicities, particularly in treatment with cytotoxic immu-
`noconjugates as envisaged in cancer patients, this is not
`desirable. An alternative approach is to devise methods for
`specific abrogation ofantibody responses along the lines already
`reported with daunomycin-antibody conjugates or by some
`other form of immune intervention.
`Considerable advances are being made in the clinical use of
`monoclonal antibodies and immunoconjugates. These include
`immunoscintigraphy with radioisotope-labelled antibodies for
`tumour detection, and monoclonal antibodies can be viewed as
`a new class of immunosuppressive agents in transplantation.
`Promising results are also being obtained in solid tumour
`therapy with immunoconjugates and one anticipates that this
`approach to cancer treatment will be even more effective with
`improvements in conjugate design.
`
`REFERENCES
`BALDWIN R.W. (1985) Design and development of drug-monoclonal
`antibody 791T/36 for cancer therapy. In: Monoclonal antibody
`therapy of human cancer (eds K. Foon and A.C. Morgan), p. 23.
`Martinus Nijoff Publishing, Boston, MA.
`BALDWIN R.W. & BnERs V.S. (1987) Monoclonal antibodies and
`immunoconjugates for cancer treatment. In: Cancer Chemotherapy
`and Biological Response Modifiers Annual 9 (eds H.M. Pinedo, D.L.
`Longo and B.A. Chabner), p. 409. Elsevier Science Publishers,
`Amsterdam.
`BALDWIN R.W. & BYERS V.S. (1988) Monoclonal antibodies in colo
`rectal cancer dia