Monoclonal Antibodies

Journal of Clinical Pharmacy and Therapeutics (1997) 22, 7–19
REVIEW ARTICLE
Monoclonal antibodies in drug targeting
R. Panchagnula MPharm PhD and C. S. Dey PhD*
Departments of Pharmaceutics and *Biotechnology, National Institute of Pharmaceutical Education and Research (NIPER),
Sector-67, S.A.S. Nagar, Punjab-160 062, India
SUMMARY
The objective of drug targeting is to deliver drugs
to a specific site of action through a carrier
system. In cancer chemotherapy, cytotoxic drugs
kill cancerous cells but also damage normal cells.
Monoclonal antibodies generated against specific
antigens, when conjugated to cytotoxic drugs, can
selectively deliver drugs to cancer cells while
minimizing damage to normal cells. Of all the
carrier systems available, monoclonal antibodies
are gaining importance because of their high
specificity. The purpose of this review is to pro-
vide a comprehensive account of the use of mono-
clonal antibodies in drug targeting, highlighting
their scope and limitations.
INTRODUCTION
The selective delivery of drugs to their site of action
should increase their therapeutic effectiveness while
minimizing unwanted side-effects (toxic effects). In
many instances, a drug has limited or no access to its
intended site of action or is rapidly metabolized or
excreted. In other instances, the drug distributes freely
throughout the body, however, it not only acts on the
desired target site (tissues) but also causes undesirable
effects on non-target tissues. With drug targeting the
drug is linked reversibly to a pharmacologically inert
and biodegradable carrier molecule. The conjugate
delivers the drug at the target site.
Drug targeting may use both passive and active
systems. In passive targeting, the distribution of the
drug–carrier complex is restricted to the capillary bed
(first-order targeting), whereas in active targeting,
selectivity in delivery of the drug–carrier complex
occurs. Such selective delivery to cells or tissues is
referred to as second-order targeting. Delivery to a
preselected intracellular organelle (e.g. lysosomes) is
known as third-order targeting (1).
Although the principle of drug targeting is simple,
the main problem is finding a carrier molecule that
delivers the drug to the target site. Another problem is
the conjugation of the drug with the carrier molecules.
Target site recognition became more practical with the
discovery that the cell surface has many receptors and
with the progress achieved in the development of
monoclonal antibodies.
The following factors and requirements are of
importance when considering the development of a
drug–monoclonal antibody complex or conjugate for
drug targeting (1–3).
- The recognition site for the monoclonal antibody
should be located on the surface of the cell.
- The antibodies should have sufficient tumour tissue
specificity.
- The extent of localization of the antibody at the
target site.
- Biodistribution of the drug–antibody conjugate in
the body relative to that of the parent antibody.
- Stability of the drug–antibody conjugate in blood.
- The host toxicity of the conjugate.
- The conjugate must be biodegradable and non-
immunogenic.
- Drugs should be released upon interaction between
the carrier molecule and the cell.
ANTIBODIES
Antigens and antibodies
Antigens (or immunogens) are defined as substances
that induce an immune response. The immune
response produced may be an antibody (humoral) or
production of sensitized cells (cellular response).
Correspondence: R. Panchagnula, Department of Pharmaceutics,
National Institute of Pharmaceutical Education and Research
(NIPER), Sector-67, S.A.S. Nagar, Punjab-160 062, India.
? 1997 Blackwell Science Ltd 7

Usually both responses are stimulated. Proteins
formed in response to an immunogen are defined
as antibodies. Antibodies are produced by B lym-
phocytes as one component of the immune response
following recognition of foreign substances (antigens),
such as bacterial or viral proteins. Under normal
conditions, antigenic stimulation results in the gener-
ation of a multiple population of sensitized cells (B
lymphocytes), each of which are ‘programmed’ to
produce an antibody to a single determinant. A single
protein may contain multiple determinants, and immu-
nization typically leads to the production of ‘poly-
clonal’ antisera containing many types of antibodies
(4). Monoclonal antibodies are antibodies produced by
clones of a single cell which recognize and bind to a
specific antigen. Theoretically, it is possible to gener-
ate a highly specific antibody against an antigen on a
particular cell type. Therefore, therapeutic agents con-
jugated to such specific antibodies should reach the
targeted cell type in high concentrations, leading to
improvement in therapeutic efficacy at much lower
concentrations than if the drug were administered in
free form. Cancer-cell specific monoclonal antibodies
have been raised successfully. Because they have high
affinity, monoclonal antibodies, immunoconjugates
(containing toxins (5–11), cytotoxic drugs (6, 12, 13)
and radioisotopes (6, 14–17)) have been investigated
extensively in cancer chemotherapy, and some of
these conjugates have reached preclinical and also
clinical trial stages in the management of colon, breast,
skin and bone cancers (10–12, 18).
Structure of antibodies
Antibodies are glycoproteins comprising 82–96%
polypeptides and 4–18% carbohydrates. Proteins with
antibody activity are generally called ‘immunoglobu-
lins’. All immunoglobulins have a common structure
of four peptide chains. The two identical long chains
are called heavy chains (H chains) and the two
identical short chains are called light chains (L chains).
These chains are held together by non-covalent and
covalent inter-chain disulphide bonds that permit
mobility. The carbohydrate portion of the immuno-
globulin molecule is covalently bonded to amino acids
in the polypeptide chains (19–21).
Proteases can cleave the molecule into two types of
functional domain. Digestion with papain cleaves the
molecule at the N-terminal side of the disulphide bond,
yielding three fragments of approximately equal size.
Two of these fragments are identical and retain the
antigen binding capacity associated with an intact
antibody. These are known as Fab fragments and are
composed of entire light chains and a portion of heavy
chains. The third fragment (known as Fc fragment) has
no antigen-binding capacity and is crystallizable. The
Fc fragment is composed of the C-terminal half of the
heavy chain and retains the other biological activities
associated with the immunoglobulin molecule (inter-
action with the complement system and tissue bind-
ing). Digestion with pepsin cleaves the antibody
molecule at the C-terminal of the disulphide bond.
This results in the F(ab)2 fragment, comprising two Fab
fragments linked by a disulphide bond. The remaining
molecule undergoes extensive degradation. The
structure of Fc fragment is the same for all of the
antibodies, but the structure of Fab fragment varies
from antibody to antibody.
Monoclonal antibody production
The logic underlying the generation of monoclonal
antibodies is deceptively simple. B lymphocytes pro-
duce antibodies of interest, but they lack a sustained
ability to grow in culture. Conversely, tumour cells,
derived from B cells, maintain long-term growth but
may not secrete antibodies. If these two types of cells
are fused, the property of long-term growth can be
conferred upon the antigen-specific B cells without
loss of antibody secretion.
Mixing two populations of cells in the presence of
compounds such as polyethylene glycol (fusogens)
leads to fusion of the cells. Subsequent fusion of the
nuclear membrane then results in the formation of a
tetraploid hybrid cell. After fusion, two types of cells
able to maintain sustained growth remain in the
culture: (a) the fusion products of tumour (myeloma)
cells and antigen-activated B cells, and (b) the original
unfused cells.
Because the original tumour cells have less DNA
to replicate and less protein to make per cell, one
might expect that they would eventually overgrow
the fusion product. To overcome this, mutant tumour
cells which are deficient in a particular enzyme, hypox-
anthine guanine phosphoribosyl transferase (HGPRT)
are generated and used for fusion. HGPRT catalyses
the reaction of hypoxanthine and guanine with
5-phosphoribosyl-1-pyrophosphate to form nucleotide
inosine-5-phosphate and guanosine 5-phosphate,
respectively. Thus, if de novo synthesis of purines is
8 R. Panchagnula and C. S. Dey
? 1997 Blackwell Science Ltd, Journal of Clinical Pharmacy and Therapeutics, 22, 7–19

blocked, this enzyme enables the cell to use hypox-
anthine and guanine to generate all the purine
monophosphates necessary for cell growth.
Addition of a reagent such as aminopterin, which
blocks de novo purine biosynthesis, will block the
growth of the unfused tumour cells because they are
unable to utilize the ‘rescue’ pathway involving
HGPRT. Moreover, the hybrids will continue to grow
because the necessary HGPRT gene product is pro-
vided by the wild-type B cell parent. However, in
order to facilitate purine biosynthesis by the alterna-
tive pathway, hypoxanthine and thymine has to be
provided in the medium (HAT selective medium).
Because unfused B cells are intrinsically short-lived, the
only cells growing out of the fusion in the presence of
aminopterin should be the fused B cell-myeloma
product. These cells produce antibodies which are
subjected to extensive screening to obtain the clone
that secretes the antibody of interest. Various articles,
books and handbooks are available that describe the
concepts and procedure of generation of monoclonal
antibodies in greater detail (22–27).
DRUG–ANTIBODY CONJUGATES
Many cytotoxic drugs have been conjugated with
monoclonal antibodies (3, 19). These conjugates have
been used to study drug localization in tumours and
modulation of drug toxicity. They have been found
to be useful in the management of various types
of carcinomas, such as colorectal, gastric, ovarian,
epidermal, endometrial, breast, lung and pancreatic
carcinoma (3, 28–31).
Monoclonal antibodies consist of many polypeptide
chains with reactive groups, for example amino, car-
boxylic and hydroxylic. These reactive groups are
important for binding with antigens. During the con-
jugation process with cytotoxic drugs, the reactive
groups should be protected, although some of the
amino, hydroxyl and carboxyl groups are utilized
for conjugating the antibodies. However, these are
made available again upon tumour localization or
intracellular release (3).
In the development of monoclonal antibodies for
cytotoxic drug targeting, the conjugation process
must not affect the cytotoxic activity of the drug and
the specificity of the monoclonal antibody while
maintaining stability in the circulation prior to reach-
ing the target site (32, 33). Although many methods
are available for general conjugation of small mol-
ecules to macromolecules, several of these are not
gentle and/or the linkage between the drug and
antibody is very unstable in vivo. Most commonly,
NH2, SH, tyrosine residues and aldehyde groups of
antibodies are involved with reactive groups of drugs
in forming drug–antibody conjugates.
Diazotization is a technique in which tyrosine
residues of an antibody participate in the conjugation
of the drug (34). This procedure is usually not suitable
for conjugating cytotoxic drugs because the reactive
tyrosine of the antibody confers specificity (35). Con-
jugation involving periodate oxidation of the amino
sugar moiety of anthracycline derivatives resulted in
loss of cytotoxicity (36).
Generally, antibodies contain large amounts of
lysine moieties which are commonly the preferred
sites for conjugation with cytotoxic drugs. In this type
of conjugation, the carboxylic group of a cytotoxic
drug is reacted with the amino group of antibody
lysine. Chlorambucil has been coupled with antibody
through the amino group of lysine, although the
drug–antibody conjugation was through the for-
mation of an ionic complex rather than covalent
bonding (37).
Coupling of the amino sugar residues of anthra-
cycline derivatives gave the best results when com-
pared to other methods involving other functional
groups of those drugs (38). Formation of a cis-aconityl
linkage between the amino sugar of anthracycline
derivatives and antibodies leads to the most stable
conjugates under physiological pH (39) and releases
the free drug in the acidic environment of lysosomes
after transportation into the cells (40). The efficacy of
these drug–antibody conjugates proved that covalent
bonding between drug and antibody does not reduce
drug activity.
Monoclonal antibodies may behave differently with
different drugs, therefore general conclusions drawn
from the above conjugation studies may be very
risky. This point has been clearly demonstrated by
the results obtained with vindesine–antibody and
arabinoside–antibody conjugates (41, 42). Therefore, it
is always better to evaluate drug–antibody conjugates
individually (33).
The number of antibody binding sites available on
each cell surface, the number of cells having antibody
binding sites, the size of the tumour, the presence
and activity of circulating tumour antigens and the
immunoreactive component of antibody are some
Monoclonal antibodies in drug targeting 9

additional factors which deserve care and consider-
ation when developing drug–monoclonal conjugates
(43, 44). Ghose and colleagues successfully conjugated
many drugs, such as chlorambucil, adriamycin, dauno-
mycin, methotrexate, trenimon, vindesine and others,
without sacrificing the cytotoxicity of drugs or speci-
ficity of antibodies (37, 45–49). All these conjugates
showed activity and effectiveness against cancer cell
lines in vitro, but were less potent than the free drug
(50–52). Loss of potency was compensated by
improved specificity of the drug antibody to the
cancer cell lines. Loss of potency was also compen-
sated in vivo by increased circulation time and longer
residence time of the drug–antibody conjugate at the
target site. However, in some cases the drug conjugate
has shown higher potency than the free drug (53).
One of the problems with drug–antibody therapy is
the low amount of drug delivered at the target site,
because only one drug molecule is conjugated to each
antibody (54). Techniques involving site-specific link-
age using spacers such as dextran and human serum
albumin provide a means for improving drug loading.
Modifying the conjugation sites on the antibody may
improve drug loading but this could decrease the
immunoreactivity of the drug–antibody conjugate
(decreased specificity). Using human serum albumin
(HSA) as a carrier results in up to 30 methotrexate
molecules conjugating to one or two molecules of
HSA. This methotrexate–HSA conjugate can then be
coupled to antibody (52). The drawback with this
procedure is the size of the drug–HSA–antibody
conjugate, leading to rapid clearance from the circu-
lation and a decrease in specificity. In another study
approximately 30–50 molecules of methotrexate were
conjugated to one molecule of anti-carcinoembryonic
antigen (CEA) monoclonal antibody using an amino–
dextran carrier (55, 56). For anthracyclines, loading of
up to 500 molecules per antibody was achieved by
using amino dextran derivatives (57, 58). This type
of conjugation is promising because there was no
significant loss of specificity.
Several cytotoxic drugs (e.g. chlorambucil (37, 53,
59, 60), trenimon (61), melphalan (60, 62), cisplatin
(63), anthracyclines (38, 55, 57, 64–69), 5-fluorouracil
(70), etc.) have been conjugated with tumour specific
antibodies and evaluated for both tumour localization
and therapeutic effect. Some of these are presently
undergoing Phase II clinical studies.
Other classes of cytotoxic drugs such as mylansoids
(71, 72) and enediyne antibiotics (73–75) have also
been conjugated to monoclonal antibodies. These
conjugates are being evaluated for specificity and
cytotoxicity against a wide range of tumours in
different tumour models.
Methotrexate conjugated to monoclonal antibody
has been compared with methotrexate-IgG (non-
specific antibody) and free methotrexate against EL4
lymphoma (48). The methotrexate–antibody conju-
gate was 3 and 7 times more effective against EL4
lymphoma than methotrexate-IgG and free metho-
trexate, respectively. Similar results were reported by
Pimm et al. (76) and Ballantyne et al. (77) using
791T/36 antibody instead of EL4 lymphoma.
Although tumour localization of methotrexate was
observed with methotrexate-IgG, achieving therapeu-
tic concentrations of methotrexate in tumour tissue
was practically impossible. In another study, a CEA
carrier system (56) was used to conjugate a mono-
clonal antibody (791T/36). Although high drug load-
ing and the therapeutic efficacy was achieved, the
studies involved experimental subcutaneous tumours
in animals and the conclusions drawn could be mis-
leading because the effects could be due to ‘reservoir
effect’ arising from local application rather than to any
targeting effect of the monoclonal antibodies. Hence,
appropriate controls are important in the evaluation
of monoclonal antibodies for targeting. In order to
increase tumour localization of methotrexate, it was
conjugated with F(ab)2 and the conjugate evaluated
against EL4 mouse lymphoma. Methotrexate–F(ab)2
conjugates were not as effective as the methotrexate–
monoclonal antibody conjugate. A methotrexate-
resistant tumour cell line was recently treated with
methotrexate–HSA–monoclonal antibody by Affleck
and Embelton (78), and in vitro results indicate that it
is possible to overcome resistance to methotrexate.
Previous reports indicated poor penetration of the
drug into tumour tissue due to rapid in vivo clearance
and the size of the conjugate. Therefore, further
evaluation of the drug conjugate is necessary to draw
firmer conclusions. Although in vivo clearance can be
reduced by using a biphasic antibody, more studies
involving size and penetration of conjugates into
tissues are required.
Vinca alkaloids
Vinca alkaloids have high molar potency, and if
potency could be retained after conjugation with a
monoclonal antibody, the dose delivered to the target

site would be sufficient. Vindesine (a vinca alkaloid)
has been conjugated with different anti-tumour anti-
bodies, such as anti-CEA, melanoma, osteosarcoma
and others (79–82). Studies with vindesine–anti-CEA
conjugates (82) indicate that they increase the thera-
peutic index of vindesine by decreasing the toxicity
and increasing the specificity to tumour. These find-
ings were substantiated by Casson et al. (83). Further-
more, evaluation of the conjugate against non-CEA
producing colon carcinoma xenograft and CEA-
producing xenograft showed that significant retar-
dation in the growth of tumour was observed only in
CEA-producing tumour (84).
IMMUNOTOXINS
Immunotoxin therapy has some distinct advantages,
for instance:
(a) The naturally occurring toxins used have very
specific biological pathways in producing their
cytocidal effects.
(b) The cytotoxic activity of the toxin that is conju-
gated to the antibody does not involve any other
secondary agent(s).
(c) Theoretically, immunotoxins should not bind to
non-malignant cells, and even if they do bind, the
internalization of the agent should not be sufficient
to neutralize the therapeutic effect.
Toxins that are used to conjugate with antibodies
are biological molecules. Diphtheria toxin (DTx) and
Pseudomonas exotoxin (PE) are obtained from bacteria.
Ricin and abrin are found in plants. The basic molecular
structure of these toxins consists of two (three in the
case of PE) polypeptide chains; the A chain (catalytic
unit) and the B chain (binding unit, and also a trans-
location domain in PE) held together by disulphide
bonds. Immunotoxin conjugates, made up of one of
these toxins, bind to the cell surface antigen via the
specific antibodies. The surface bound toxin becomes
internalized along with the antibody. In the endosomal
compartment, the toxin undergoes processing and the
catalytic domain of the toxin is released into the
cytosol of the targeted cells, thereby inhibiting protein
synthesis and killing the tumour cells (10, 11, 85–89).
If the antibody conjugated to the whole toxin is of
very high potency it has the potential for cross-
reacting with cells that are devoid of target antigens to
produce severe non-specific toxicity. There are several
examples of toxicity observed both in laboratory tests
and in clinical trials (11). To eliminate this problem, the
A chain is enzymatically separated from the toxin
before being conjugated with the antibody. The anti-
body provides binding capacity, and the A chain,
being catalytic, imparts cytotoxic effect(s). This
approach has led to several immunotoxins which have
been shown to be active in vitro. Some have reached
Phase I clinical trial in B cell chronic lymphocytic
leukaemia, pulmonary, colon, breast and lymph node
metastatic melanoma (10, 11, 90–93). In general, major
problems with these immunotoxins include undesir-
able humoral immune responses, instability and
reduced therapeutic efficacy due to the large size of the
immunotoxin. Immunotoxins prepared by conjugating
A chain with the antibody are poorly cytotoxic (e.g.
ricin-A chain is 100 000-fold less active) because of
poor binding capacity to the cell surface. The cyto-
toxicity of an immunotoxin can be enhanced by
restoring B chain integrity within the conjugate, but
the B chain has to be structurally altered to reduce its
galactose binding site in order to decrease non-specific
binding. Unfortunately, the cleaved toxin has always
shown reduced activity when compared to the entire
toxin. Genetic engineering of ricin to eliminate the
galactose binding sites has not been successful because
its extreme toxicity prevents successful expression in
eukaryotic systems. Ricin A and B chains have been
separately cleaved and expressed in bacteria in bio-
logically active forms and recombinant B chain with
no galactose binding activity has been generated
by substitution of a single amino acid residue (94).
Univalent Fab fragments generated by proteolytic
cleavage have shown some promise in certain cases
(95). But the avidity of the univalent fragments is less.
Moreover, absence of an Fc portion, which is respon-
sible for IgG catabolism, reduces the half-life of the
immunotoxin (10, 11).
Another group of natural toxins, trichothecene
mycotoxins, which are fungal metabolites of Fungi
imperfecti and are potent inhibitors of protein syn-
thesis, have been found to be as active as PE and a
100-fold more active than doxorubicin (2). These
trichothecene toxins were found to be highly active in
murine leukaemia, but have caused severe toxicity in
animal studies and in clinical trial. Preliminary studies
with trichothecenes conjugated to an antibody specific
to murine EL-4 thymoma have shown selective cyto-
toxicity of the toxin towards the cell in vitro as well as
in vivo (2).
Immunotoxins of bacterial, plant or fungal origin
have intrinsic cytotoxicity against normal cells and

efforts are being made towards finding better alterna-
tives. The first synthetic immunotoxin contained
fungal glucose oxidase which killed Hela cells. A
cytocidal enzyme immunotoxin was devised using
two sequentially acting enzymes, glucose oxidase and
lactoperoxidase, which kill murine plasma cytoma cells
in vitro by generating H2O2 and I2 (and/or its halo-
genated derivatives) (96). Enzymic activation of a less
cytotoxic prodrug in the vicinity of the target cells has
been demonstrated to lead to increased therapeutic
efficacy. An example is the enhanced effect of
antibody–alkaline phosphatase conjugate of etoposide
phosphate both in vitro and in vivo (97).
Antibodies conjugated to radionucleides are also
used for immunotherapy. Complete remission has
been achieved in some selected patients (15). Radio-
labelled antibody was found to be selectively localized
in lymphatic tissues (14–17). The F(ab)2 fragments
were concentrated at the site when compared to the
whole radiolabelled antibody (almost a 144-fold
increase) (15). This selectivity demonstrates the poten-
tial for monoclonal antibodies to deliver toxins (e.g.
radioisotope) to tumours in vivo.
TOXICITY AND ANTIBODIES
Although exogenous antibodies have been used for
therapeutic purposes for many years, the use of
specific antibodies to reverse toxic effects of drugs is
more recent (98). Digitalis intoxication is one of the
most common forms of toxicity reported in clinical
medicine. Treatment of digitalis intoxication is limited
by the absence of a specific antagonist. The need to
improve management of digitalis toxicity stimulated
interest in finding a specific means for reversing the
effect of this particular class of drugs. Antibodies to
digoxin and digitoxin (digitalis glycosides) neutralize
the effect of these drugs by binding to the free drug
in the blood and thus preventing them from binding
to digitalis receptors in cardiac tissues. Successful
treatment of digitalis-overdosed patients with digoxin-
specific antibody fragments has been reported (99–
101). Administration of anti-digoxin Fab fragment
reverses the digitalis effect. The total serum digoxin
concentration measured was actually higher than prior
to antibody administration. Essentially, all of the dig-
oxin was bound to antibody fragments and therefore
rendered inactive. The rise in digoxin concentration
suggests that some digoxin is extracted from tissues
(102, 103). Treatment and management of life threat-
ening digoxin overdose is now by administration of
digoxin-specific Fab fragments (104). Treatment of
drug intoxications (overdose) using i.v. administration
of monoclonal antibodies or antibody fragments is
described extensively by Sabouraud and Scherrmann
(105).
Preliminary reports suggest that Fab fragments may
be useful in treating toxicity of colchicine (106),
paraquat (107), phencyclidine (108) and tricyclic
antidepressants (109–112).
SECOND GENERATION ANTIBODIES
Second generation antibodies (chimeric antibodies) are
being developed by novel biotechnological methods
in order to improve penetration of conjugates into
tissue. Recent developments include expression of
chimeric antibodies in yeast or bacteria, production of
human monoclonal antibodies, generation of fusion
proteins, single domain antibodies and humanized
monoclonal antibodies (113, 114). A very promising
chimeric antibody is made up of mouse–human anti-
bodies in which the variable region of a mouse
monoclonal antibody is linked to human IgG1 con-
stant regions. These chimeras have been found to be
better than the parent mouse antibody in cell-mediated
cell killing assays (115, 116). Olsson and Kaplan (117)
reported the production of human–human monoclonal
antibodies of defined specificity.
Because it is still not feasible to obtain antibodies
raised against human volunteers and patients,
resources have been diverted towards humanizing the
large number of well-characterized rodent monoclonal
antibodies, including those that are specific against
human antigens, by protein and genetic engineering.
As a result, antibodies such as anti-CDN52 have
been developed and shown to be clinically active in
rheumatoid arthritis (118). Various other antibodies
have been constructed against a wide range of patho-
genic bacteria, virus and human cell surface markers
including tumour cell antigens. Some humanized
monoclonal antibodies have been conjugated with
drugs and are undergoing clinical trials (66, 119–121).
It is now possible to transplant the complimentary-
determining regions (CDRs) from a murine antibody
into a human framework (122). CAMPATH-1, a CDR
graft of anti-lymphoma monoclonal antibody has been
very successful in the therapy of non-Hodgkin’s

lymphoma and rheumatoid arthritis (123–125).
Borrebaeck et al. (126) described an in vitro method for
immunization that produces human–human hybrid-
oma secreted monoclonal antibodies specific for dig-
oxin, haemocyanine and one recombinant fragment of
gp120 envelope glycoprotein of HIV. Recent devel-
opments in humanized antibodies have been reviewed
by Winter and Harris (114).
With Staphylococcus aureus, a gene for a nuclease
has been joined with a mouse heavy chain gene to
produce a recombinant fusion protein immunotoxin
(127). Fusion protein tissue type plasminogen
activator has been used in removing blood clots.
Fibrin-specific monoclonal antibodies were coupled to
urokinase (128), or a recombinant fusion protein was
made by fusing appropriate domains of fibrin-specific
monoclonal antibody with DNA sequences coding for
the plasminogen activator function (129). Human
immunodeficiency virus (HIV) through its surface coat
protein gp120 binds to CD4 molecules present on the
surface of helper T-lymphocytes, thereby acting as a
cause of acquired immunodeficiency syndrome (AIDS).
CD4 immunoadhesin binds to gp120 and blocks HIV.
It has a long plasma half-life and is capable of binding
to Fc receptors. Recombinant CD4 immunoadhesin
modulates antibody-dependent cell-mediated cyto-
toxicity towards HIV infected cells (130, 131). In
addition, CD4 immunoadhesin permeates efficiently
through the placenta (113) thereby making possible
perinatal treatment of HIV infection.
Another potentially useful biotechnological
approach is the generation of a bispecific F(ab)2
fragment comprising two Fab fragments, one cleaved
from an anti-CD3 monoclonal antibody and the other
from an anti-P-glycoprotein (the protein responsible
for the phenomenon of multiple drug resistance in
drug resistant cancer cells) monoclonal antibody (132,
133). This bispecific antibody (F(ab)2) enhanced the
binding of the antibody and increased the cytotoxicity
of human peripheral blood mononuclear cells to
P-glycoprotein-positive kidney cancer cells (132).
Using a similar approach, heterobifunctional antibody
duplexes have been prepared in which one antibody
recognizes one element on the tumour cell while the
other antibody binds specifically to antigen–receptor
complexes on T-cells. These in turn become activated
and lyse the tumour cells (134). As presented at the
Fourth International Conference on ‘Bispecific Anti-
bodies and Cellular Toxicity’ (held in Florida, U.S.A.,
1–5 March 1995), several bispecific antibodies such as
MDX-210, MDX-240, 2B1 etc. are at various stages of
clinical trials (135), sponsored by large biotechnology
companies such as Genentech, Scotgen Pharmaceuti-
cals, Protein Design Labs, Centecor and ImmunoGen.
Other biotechnology companies are actively partici-
pating in antibody development programmes. Some of
these bispecific antibodies are being used as carriers for
cytotoxic agents such as scoparin, vinca alkaloids,
methotrexate, ricin-A chain and alkaline phosphatase
(136). Bispecific antibodies are also used for radio-
imaging and radioimmuno surgery in CEA positive
tumours in vivo and therapeutic delivery of 10
boron to
melanomas and glioblastomas (135). Moreover, bi-
specific antibodies have been successfully tested
in vitro as a vehicle for vaccination of tetanus toxoid
(135). Bispecific antibodies have been shown to be
effective in humans.
Development of another kind of second generation
antibody, the single domain antibody, is also in
progress. The VH domain of the antibody heavy chain
is found to bind antigen with high affinity in the
absence of the VL light chain domains (137). Consider-
able amounts of VH can be obtained from bacteria
(138). By chain shuffling of either mutated or non-
mutated V genes, selection of high-affinity monoclonal
antibodies of VH as well as VL domains can be
generated (2, 139). These VH domains may serve as an
alternative to monoclonal antibodies. The small size of
VH- or VL-conjugated toxins may prove to be very
useful if there is no lack of specificity (140).
LIMITATIONS AND CONCLUSIONS
Drug–monoclonal antibody conjugates are currently
undergoing clinical trials (Phase II). Much of the
literature available indicates that ‘conjugation’ of
drugs with antibodies does not necessarily lead to loss
of activity. One successful application is in the treat-
ment of drug overdose using drug-specific Fab frag-
ments. Other effective applications involve the use of
second generation monoclonal antibodies, especially
bispecific and humanized antibodies, cancer imaging
(111
ln labelled murine antimyosin monoclonal anti-
body for imaging of myocardial damage; approved
for clinical use (141)) and monoclonal antibodies as
experimental probes.
Over the last two decades many cytotoxic drugs
have been used for targeting cancer tissues. There
are limitations in spite of the increasing success of

antibody therapy. Conjugates administered intra-
venously may not reach the target sites because of
non-specific uptake and catabolism, immunocross-
reactivity of the antibody with normal tissue antigens
and inadequate diffusion of the antibody into the
tumour interstitium. The use of native antibodies is
determined by whether they efficiently trigger the
destruction of the target, or whether they have
effective blocking or neutralizing activity. Further-
more, when an animal monoclonal antibody is injected
into a human patient, it may be recognized as a foreign
substance and may stimulate the patient’s immune
system to make antibodies which react with the
therapeutic monoclonal antibodies. The use of drug–
antibody conjugate becomes all the more difficult if it
is toxic. A further problem arises when antibody
binding to a target molecule induces the disappearance
or alters the surface antigens of the target cell.
Antibodies carry a signal in the Fc domain for their
uptake by the macrophage-like cells in the reticulo-
endothelial system. The function of these cells is to
ingest foreign antigens. If these cells ingest drug–
antibody conjugates, then they are killed along with
the target tissue, leading to the destruction of an
important part of the natural immune system.
Although size and specificity of drug-conjugates
may be approached through second generation anti-
bodies, a major hurdle is the non-availability of human
monoclonal antibodies. Once the human monoclonal
antibody is available, limitations associated with
specificity and immunogenicity of drug–antibody
conjugates should be markedly decreased.
Basic research in the area of genetic engineering and
cell biology of receptor internalization and signalling
is essential for developing antibodies which are not
taken up by reticuloendothelial cells. Results of numer-
ous studies examining the localization and anti-tumour
effectiveness of monoclonal antibodies, especially bi-
specific and humanized antibodies conjugated with
cytotoxic agents should appear soon. The use of
radiolabelled monoclonal antibodies for radioimaging
should provide additional information on antibody
specificity in vivo. Antibody therapy should extend the
armamentarium of clinicians to combat disease in
the not too distant future.
REFERENCES
1. Friend DR, Pangburn S. (1987) Site specific drug
delivery. Medical Research Reviews, 7, 53–106.
2. Hinman LM, Yarranton G. (1993) New approaches to
non-immunogenic antibody cancer therapies. Annual
Reports in Medicinal Chemistry, 28, 237–246.
3. Pimm MV. (1988) Drug-monoclonal antibody conju-
gates for cancer therapy: Potentials and limitations.
CRC Critical Reviews in Therapeutic Drug Carrier
Systems, 5, 189–227.
4. Baldwin RW. (1983) Monoclonal antibodies for drug
targeting in cancer therapy. Pharmacy International, 4,
137–141.
5. Derbyshire EJ, Wawrzynczak EJ. (1992) An anti-mucin
immunotoxin BrE-3-Ricin A chain is potently and
selectively toxic to human small-cell lung cancer.
International Journal of Cancer, 52, 624–630.
6. Dillman RO, Johnson DE, Shawler DL. (1988) Compari-
sons of drugs and immunoconjugates. Antibody Im-
munoconjugates and Radiopharmaceuticals, 1, 65–77.
7. Engert A, Barrowa F, Jung N, et al. (1990) Evaluation
of Ricin A chain containing immunotoxins directed
against the CD30 antigens as potential reagents for the
treatment of Hodgkin’s disease. Cancer Research, 50,
84–88.
8. Engert A, Martin G, Pfreundschuh M, et al. (1990)
Antitumor effects of Ricin A chain immunotoxins
prepared from intact antibodies and Fab fragments in
solid human Hodgkin’s disease tumors in mice. Cancer
Research, 50, 2929–2935.
9. Vitetta ES, Fulton RJ, May RD, Till M, Uhr JN. (1987)
Redesigning nature in poison to create anti-tumor
reagents. Science, 238, 1098–1104.
10. Vitetta ES, Thorpe PE, Uhr JN. (1993) Immunotoxin:
magic bullets or misguided missiles. Trends in Pharma-
cological Sciences, 14, 148–154.
11. Wawrzynczak EJ. (1991) Systemic immunotoxin
therapy of cancer: advances and prospects. British
Journal of Cancer, 64, 624–630.
12. Pietersz GA, Smith MJ, Kanells J, Cunningham Z,
Sacks NPM, McKenzie IFC. (1988) Preclinical and
clinical studies with a variety of immunoconjugates.
Antibody Immunoconjugates and Radiopharmaceuticals, 1,
79–103.
13. Spitler L, delRio M, Khentigan A, et al. (1987) Therapy
of patients with malignant melanoma using a mono-
clonal antimelanoma antibody Ricin A chain. Cancer
Research, 47, 1717–1723.
14. McGargkey C. (1974) Feasibility of tumor immuno-
therapy using radioiodinated antibodies to tumor-
specific cell membrane antigens with emphasis on
leukemias and early metastases. Oncology, 29, 302–307.
15. Ballau B, Levine G, Hakala TR, Solter E. (1979) Tumor
location detected with radioactively labeled mono-
clonal antibody and external scintigraphy. Science, 206,
844–846.

16. Order SL, Sleeper AM, Stillwagon GB, Klien JL, Leicher
PK. (1990) Radiolabeled antibodies and potential cancer
therapy. Cancer Research, 50 (Suppl.), 1011–1013.
17. Goldenberg DM, Larsom SM, Reisfeld RA, Schlom J.
(1995) Targeting cancer with radiolabeled antibodies.
Immunology Today, 16, 261–264.
18. Thomas H. (1992) New horizons in cancer therapy.
Drugs of Today, 28, 311–331.
19. Brodsky FM. (1988) Monoclonal antibodies as magic
bullets. Pharmaceutical Research, 5, 1–9.
20. Ganong WF. (1985) Review of Medical Physiology.
pp. 426–427. Lange Medical Publications, Los Altos,
CA.
21. Goding JW. (1983) Monoclonal Antibodies: Principles and
Practice. pp. 7–10. Academic Press, New York.
22. Catty D. (1986) Antibodies Volume 1. A Practical
Approach. pp. 6–17. IRL Press, Washington DC.
23. Harlow E, Lane D. (1988) Antibodies: A Laboratory
Manual. Cold Spring Harbor Laboratory, New York.
24. Kohler G, Milstein C. (1975) Continuous cultures of
fused cells secreting antibody of predefined specificity.
Nature, 256, 495–497.
25. Milstein C. (1980) Monoclonal antibodies. Scientific
America, 243, 66–74.
26. Roitt IM. (1987) Essential Immunology. Blackwell
Scientific Publications, Oxford.
27. Zola H. (1987) Monoclonal Antibodies: A Manual of
Techniques. pp. 1–11. CRC Press Inc., Baco Raton, FL.
28. Herlyn M, Stepleweski Z, Herlyn D, Koprowski G.
(1979) Colorectal carcinoma-specific antigen detection
by means of monoclonal antibodies. Proceedings of the
National Academy of Sciences of the USA, 76, 1438–
1442.
29. Hockey MS, Stokes HJ, Thomson H, et al. (1984)
Carcinoembryonic antigen (CEA) expression and
heterogenicity in primary and autologous metastatic
gastric tumors demonstrated by monoclonal antibodies.
British Journal of Cancer, 49, 192–233.
30. Kennett RH, Gilbert F. (1979) Hybrid myelomas pro-
ducing antibodies against a human neuroblastoma anti-
gen present on fetal brain. Science, 203, 1120–1122.
31. Koprowaski G, Stepleweski Z, Herlyn D, Herlyn M.
(1978) Study of antibodies against human melanoma
produced by somatic cell hybrids. Proceedings of the
3409.
32. Rowland GF. (1983) The use of antibodies and polymer
conjugates in drug targeting and synergy. In: Targeted
Drugs. ed Goldberg E, pp. 57–72, John Wiley, New
York.
33. Upeslacis J, Hinman L. (1988) Chemical modification of
antibodies for cancer chemotherapy. Annual Reports in
Medicinal Chemistry, 23, 151–160.
34. Mathe G, Loc TB, Bernard H. (1958) Effet sur la
leucemie L1201 de la souris d’une combinaison par
diszotation d’A-methopterine et de gamma-globulines
de hamsters porteur de cette leucemie par hereogreffe.
Comptes Rendues, 246, 1626–1628.
35. O’Neill GJ. (1979) The use of antibodies as drug
carriers. In: Drug Carriers in Biology and Medicine. ed
Gregoriadis G, pp. 23–41, Academic Press, London.
36. Hurwitz E, Levy R, Maron R, Wilchek M, Arnon R, Sela
M. (1975) The covalent bonding of daunomycin and
adriamycin to antibodies with retention of both drug
and antibody activities. Cancer Research, 35, 1175–
1181.
37. Ghose T, Norwell S, Guclu A, Cameron D, Bodurtha A,
Macdonalds AS. (1972) Immunotherapy of cancer with
chlorambucil carrying antibody. British Medical Journal,
3, 495–499.
38. Gallego J, Price MR, Baldwin RW. (1984) Preparation
of four daunomycin–monoclonal antibody 791T/36
conjugates with anti-tumor activity. International Journal
of Cancer, 33, 737–744.
39. Yang HM, Reisfeld RA. (1988) Doxorubicin conjugated
with monoclonal antibody directed to human
melanoma-associated proteoglycan suppresses the
growth of established tumor xenografts in nude mice.
Proceedings of the National Academy of Sciences of the
USA, 85, 1189–1193.
40. Rowland GF. (1983) Use of antibodies target drugs to
tumor cells. Clinics in Allergy and Immunology, 3,
235–257.
41. Arnon R, Hurwitz E. (1985) Monoclonal antibodies as
carriers for immuno-targeting of drugs. In: Monoclonal
Antibodies for Cancer Detection and Therapy. eds Baldwin
RW, Bayer VS, pp. 367–383, Academic Press, London.
42. Worren W. (1992) Coupling of monoclonal antibodies.
In: Monoclonal Antibodies. eds Peters JH, Baumgarten H,
pp. 285–291, Springer-Verlag, Berlin.
43. Schlom J, Weeks MO. (1984) Potential clinical utility of
monoclonal antibodies in the management of human
carcinomas. In: Important Advances in Oncology. eds
DeVita V, Halman S, Rosenberg S, pp. 170–192, J.B.
Lippincot, Philadelphia.
44. Schlom J. (1988) Monoclonal antibodies in cancer
therapy: the present and future. Biopharmacy, 9 (Sept.),
44–48; see also Pharmaceutical Technology, 88 (Sept.),
56–60.
45. Ghose T, Blair AH. (1978) Antibody-linked cytotoxic
agents in the treatment of cancer. Journal of the National
Cancer Institute, 61, 657–676.
46. Ghose T, Blair AH, Kulkarni P, Vaughan K, Norvell S,
Beltitsky P. (1982) Targeting of radionucleotides and
drugs for the diagnosis and treatment of cancer. In:
NATO Advanced Study Institute Series A. Plenum Press,
New York. Life Sciences, 47, 55–82.

47. Ghose T, Blair AH, Kulkarni P. (1983) Preparation of
antibody-linked cytotoxic agents. Methods in Enzy-
mology, 93, 280–283.
48. Ghose T, Blair AH, Uadia P, et al. (1985) Antibodies as
carriers of cancer chemotherapeutic agents. Annals of
New York Academy of Sciences, 446, 213–227.
49. Kulkarni PJ, Blair AH, Gosh T, Mammen M. (1985)
Conjugation of methotrexate to IgG antibodies and
their F(ab*)2 fragments and the effect of conjugate on
tumor growth in vivo. Cancer Immunology Immuno-
therapy, 19, 211–216.
50. Embelton MJ. (1986) Targeting of anti-cancer therapeu-
tic agents by monoclonal antibodies. Biochemical Society
Transactions, 14, 393–396.
51. Endo N, Taketda Y, Kishida K, et al. (1987) Target
selective cytotoxicity of methotrexate conjugates with
monoclonal anti-MM46 antibody. Cancer Immunology
Immunotherapy, 25, 1–6.
52. Garnett MC, Embelton MJ, Jacobs E, Baldwin RW.
(1983) Preparation and properties of a drug-carrier–
antibody conjugate showing selective antibody-
directed cytotoxicity in vitro. International Journal of
Cancer, 31, 661–670.
53. Smyth MJ, Pietersz GA, Classon BJ, McKenzie IFC.
(1986) Specific targeting of chlorambucil to tumor with
the use of monoclonal antibodies. Journal of the National
Cancer Institute, 76, 503–540.
54. Smyth MJ, Pietersz GA, McKenzie IFC. (1987) Specific
targeting of chlorambucil to tumor with the use of
monoclonal antibodies. Cancer Research, 47, 62–67.
55. Hurwitz E, Schechter B, Arnon R, Sela M. (1979)
Binding of anti-tumor immunoglobulins and their
daunomycin conjugates to the tumor metastases, in
vitro and in vivo studies with Lewis lung carcinoma.
International Journal of Cancer, 24, 461–464.
56. Shih LB, Sharkey RM, Primus FJ, Goldberg DM. (1988)
Site-specific linkage of methotrexate to monoclonal
antibodies using an intermediate carrier. International
Journal of Cancer, 41, 832–839.
57. Tsukada Y, Ohkawa K, Hibi N. (1987) Therapeutic
effect of treatment with polyclonal or monoclonal
antibodies to á-fetoprotein that have been conjugated
to daunomycin via dextran bridge: studies with an
á-fetoprotein-producing rat hepatoma tumor model.
Cancer Research, 47, 4293–4295.
58. Arnon R. (1982) Targeting of Drugs. eds Gregoriadis G,
Senior J, Trouet A, p. 31, Plenum Press, New York.
59. Tai J, Blair AH, Ghose T. (1979) Tumor inhibition by
chlorambucil covalently linked antitumor globulin.
European Journal of Cancer, 15, 1357–1361.
60. Smyth MJ, Pietersz GA, McKenzie IFC. (1987) The in
vitro and in vivo anti-tumor activity of N-AcMEL-
(Fab*)2 conjugates. British Journal of Cancer, 55, 7–11.
61. Ghose T, Guclu A, Raman RR, Blair AH. (1982)
Inhibition of mouse hepatoma by alkylating agent
trenimon linked to immunoglobulins. Cancer Immunol-
ogy Immunotherapy, 13, 185–189.
62. Rowland AJ, McKenzie IF, Pietersz GA. (1993)
Enhanced antitumor effects using a combination of two
antibodies conjugated to two different drugs. Journal of
Drug Targeting, 2, 113–121.
63. Schechter B, Pauzner R, Arnon R, Haimovich J,
Wilcheck M. (1987) Selective cytotoxicity against
tumor cells by cisplatin complexed to antitumor anti-
bodies via carboxymetheyl dextran. Cancer Immunology
64. Isreal M, Modest EJ, Frei E. (1975) N-Trifluoro-
acetyl adriamycin 14-veleraten an analog with greater
experimental antitumor activity and less toxicity than
adriamycin. Cancer Research, 35, 1365–1368.
65. Pietersz GA, Smyth MJ, MaKenzie TFC. (1988) The
use of anthracycline–antibody complexes for specific
antitumor activity. In: Antibody-Mediated Delivery
Systems. ed Rodwell JD, pp. 25–53, Marcel Dekker,
Basel.
66. Pimm MV, Jones JA, Price MR, Middle JG, Embleton
MJ, Baldwin RW. (1982) Tumor localization of mono-
clonal antibody against a rat mammary carcinoma
and suppression of tumor growth with adriamycin–
antibody conjugates. Cancer Immunology Immuno-
therapy, 12, 125–129.
67. Tsukada Y, Kato Y, Takeda Y, Hara T, Hirai H. (1984)
An anti á-fetoprotein antibody–daunorubicin conjugate
with a novel poly-L-glutamic acid derivative as an
intermediate drug carrier. Journal of the National Cancer
Institute, 73, 721–728.
68. Trail PA, Willner D, Lasch SJ, et al. (1993) Cure
of xenografted carcinomas by BR-96-doxorubicin
immunoconjugates. Science, 261, 212–215.
69. Young RC, Ozols RF, Myers CE. (1981) The anthra-
cycline antineoplastic drugs. New England Journal of
Medicine, 305, 139–153.
70. Hurwitz E, Kashi R, Arnon R, Wilcheck M, Sela M.
(1985) The covalent linking of two nucleotide ana-
logues to antibodies. Journal Medicinal Chemistry, 28,
137–140.
71. Chari RV, Martell BA, Gross JL, et al. (1992) Immuno-
conjugates containing novel maylansinoids: Promising
anticancer drugs. Cancer Research, 52, 127–131.
72. Shah SA, Ferris CA, Derr SM, et al. (1993) Anti-B4-
blocked ricin immunotoxin shows therapeutic efficacy
in four SCID mouse tumor model. Cancer Research, 56,
1360–1367.
73. Walker S, Landovitz R, Dong WM, Ellestad G, Kahne
D. (1992) Cleavage behavior of calicheamicin ã1
and
calicheamicin T. Proceedings of the National Academy of
Sciences of the USA, 89, 4608–4612.

74. Zein N, Sinha AM, McGahren, Ellestad GA. (1988)
Calicheamicin ã1
1: an antitumor antibiotic that cleaves
double strand DNA site specific. Science, 240, 1198–
1201.
75. Zein N, Poncin M, Nilakantan R, Ellestad GA. (1989)
Calicheamicin ã1
1 and DNA: molecular recognition
process responsible for site-specificity. Science, 244,
697–699.
76. Pimm MV, Clegg JA, Garnett MC, Baldwin RW. (1988)
Biodistribution and tumor localization of methotrexate–
monoclonal antibody 791T/36 conjugate in nude mice
with human tumor xenografts. International Journal of
Cancer, 41, 886–891.
77. Ballantyne KC, Perkins AC, Pimm MV, et al. (1988)
Biodistribution of monoclonal antibody–methotrexate
conjugate (791T/36-MTX) in patients with colorectal
cancer. International Journal of Cancer, (Suppl. 2), 103–
108.
78. Affleck K, Embleton MJ. (1992) Monoclonal antibody
targeting of methotrexate (MTX) against MTX-
resistant tumor cell lines. British Journal of Cancer, 65,
834–844.
79. Philpott GW, Gass EH, Panker CW. (1979) Affinity
cytotoxicity with an alcohol dehydrogenase–antibody
conjugate and allyl alcohol. Cancer Research, 39, 2084–
2087.
80. Rowland GF, Corvalan JRF, Axton CA, et al.
(1984) Suppression of growth of human colorectal
tumor in nude mice by vindesine–monoclonal anti-
CEA conjugates. Protides of Biological Fluids, 31, 783–
786.
81. Rowland GF, Axton CA, Baldwin RW, et al. (1985)
Antitumor properties of vindesine–monoclonal anti-
body conjugates. Cancer Immunology Immunotherapy,
19, 1–7.
82. Rowland GF, Simmonds RG, Grove VA, Mardsen CH,
Smith W. (1986) Drug localization and growth inhi-
bition studies of vindesine–monoclonal anti-CEA
conjugates in human xenograft. Cancer Immunology
83. Casson AG, Ford CHJ, Mardsen CH, Gallant ME,
Bartlett SE. (1987) Efficacy and selectivity of vindesine
monoclonal anti-carcinoembryonic antigen antibody
conjugates on human tumor cell lines grow as
xenograft in nude mice. National Cancer Institute Mono-
graphs, 3, 117–124.
84. Smith TW. (1985) Antitumor properties of vindesine–
monoclonal antibody conjugates. Cancer Immunology
85. Ghetice V, Vitetta E. (1994) Immunotoxins in therapy
of cancer: From bench to clinic. Pharmacology and
Therapeutics, 63, 209–234.
86. Ribi HO, Ludwig DS, Mercer KL, Schoolnik GK,
Kornberg RD. (1988) Three-dimensional structural of
cholera toxin penetrating a lipid membrane. Science,
239, 1272–1276.
87. Pastan I, Willingham MC, Fitzgerald DJP. (1986)
Immunotoxins. Cell, 48, 641–648.
88. Olsnes S, Moskang JO, Stenmerk H, Sandvig K. (1988)
Diphtheria toxin entry: protein translocation in the
reverse direction. Trends in Biological Sciences, 13,
348–351.
89. Clemens M. (1984) Enzymes and toxins that regulate
protein synthesis. Nature, 310, 727–729.
90. Byers VS, Rodvin R, Grant H. (1989) Phase I study of
monoclonal antibody Ricin A chain immunotoxin
xomazyme in 791 patients with metastatic colon
cancer. Cancer Research, 49, 6153–6158.
91. Hertler AA, Frankel AE. (1989) Immunotoxins, a
clinical review of their use in the treatment of malig-
nancies. Journal of Clinical Oncology, 7, 1932–1939.
92. Von Wussow P, Spitker L, Blach B, Deicher H.
(1988) Immunotherapy in patients with advanced
malignant melanoma using a monoclonal anti-
melanoma antibody Ricin A chain immunotoxin.
European Journal of Cancer and Clinical Oncology, 24
(Suppl. 2), 69–74.
93. Weiner LM, O’Dwyer J, Kitson J. (1989) Phase I
re-evaluation of an anti-breast carcinoma monoclonal
antibody 260F9-recombinant Ricin A chain immuno-
conjugate. Cancer Research, 49, 4062–4066.
94. Vitetta ES, Yen N. (1990) Expression of functional
properties of genetically engineering Ricin B chain
lacking galactose binding ability. Biochemica et Bio-
physica Acta, 1049, 151–155.
95. Chaudhary VK, Galeo MG, Fitzgerald DJ, Pastan I.
(1990) A recombinant single chain immunotoxin com-
posed of anti-Tac variable regions and a truncated
diphtheria toxin. Proceedings of the National Academy of
96. Stanislawski M, Rousseau V, Goavee M, Ito H. (1989)
Immunotoxins containing glucose oxidase and lac-
toperoxidase with tumoricidal properties: In vitro
killing effectiveness in a mouse plasmacytoma model.
97. Senter PD, Saulnier MG, Schreiber GJ, et al. (1988)
Antitumor effects of antibody–alkaline phosphatase
conjugates in combination with etoposide phosphate.
USA, 85, 4842–4846.
98. Colburn WA. (1980) Specific antibodies and Fab
fragments to alter pharmacokinetics and reverse the
pharmacologic/toxicologic effects of drugs. Drug
Metabolism Reviews, 11, 223–262.
99. Smith TW, Haber E, Yeatman L, Butler  V. (1976)
Reversal of advanced digoxin intoxication with Fab
fragments of digoxin specific antibodies. New England
Journal of Medicine, 294, 797–800.

100. Smith TW, Butler  V, Haber F, Fozzard H,
Marcus FI. (1982) Treatment of life threatening digi-
talis intoxication with digoxin-specific Fab antibody
fragments. New England Journal of Medicine, 307,
1357–1362.
101. Wenger TL, Butler VP, Haber E, Smith TW. (1985)
Treatment of 63 severely digitalis-toxic patients
with digoxin-specific antibody fragments. Journal
of American Colleges of Cardiology, 5 (Abstract),
118–123.
102. Schwartz A, Lindenmayer GE, Allen JC. (1975) The
sodium–potassium adenosine triphosphate: pharmaco-
logical, physiological and biochemical aspects. Pharma-
cological Reviews, 27, 3–134.
103. Ochs HR, Smith TW. (1977) Reversal of advanced
digitoxin toxicity and modification of pharmacokinet-
ics by specific antibodies and Fab fragments. Journal of
Clinical Investigation, 60, 1303–1313.
104. Antman EM, Wenger TL, Butler  VP, Haber E,
Smith TW. (1990) Treatment of 150 cases of life-
threatening digitalis intoxication with digoxin specific
Fab antibody fragments: Final report of a multi center
report. Circulation, 81, 1744–1752.
105. Sabouraud A, Scherrmann JM. (1993) Immunotherapy
of drug intoxications. Therapie, 49, 41–48.
106. Urtizberea M, Sabouraud A, Grandgeorge M,
Rouzious JM, Scherrmann JM. (1991) Colchicine tox-
icity by colchicine specific Fab fragments. Toxicology
Letters, 58, 193–198.
107. Chen N, Bowles MR, Pond SM. (1994) Prevention of
paraquat in suspension of alveolar type II cell by
paraquat-specific antibodies. Human Experimental
Toxicology, 3, 551–557.
108. Owens SM, Mayersohn M. (1986) Phencyclidine-
specific Fab fragments alter phencyclidine disposition
in dogs. Drug Metabolism and Disposition, 14, 52–58.
109. Brunn GJ, Keyler DE, Ross CA, Pond SM, Pentel PR.
(1991) Drug specific F(ab*)2 fragment reduces imi-
pramine cardiotoxicity in rats. International Journal of
Immunopharmacology, 13, 841–851.
110. Brunn GJ, Keyler DE, Pond SM, Pentel PR. (1992)
Reversal desimipramine toxicity in rats using drug-
specific antibody Fab* fragments. Effects of hypoten-
sion and interaction with sodium bicarbonate. Journal
of Pharmacology and Experimental Therapeutics, 260,
1392–1399.
111. Pentel PR, Ross CA, Landon J, Sidki A, Shelver WL,
Keyler DE. (1994) Reversal of desimipramine in rats
using polyclonal-drug specific antibody fragments.
Journal of Laboratory and Clinical Medicine, 123, 387–
393.
112. Keyler DE, Le Couteur DG, Pond SM, Peter ST, Pentel
PR. (1995) Effects of specific antibody Fab fragments
on desimipramine pharmacokinetics in rat in vivo and
in isolated, perfused liver. Journal of Pharmacology and
Experimental Therapeutics, 272, 1117–1123.
113. Hajela K, Galzie Z. (1991) Designer antibodies. Bio-
technological Education, 2, 154–161.
114. Winter G, Harris WJ. (1993) Humanized antibodies.
Trends in Pharmacological Sciences, 14, 139–143.
115. Nishoka Y, Sone S, Heike Y, et al. (1992) Effector cell
analysis of human multidrug resistant cell killing
by mouse–human chimeric antibody against P-
glycoprotein. Japanese Journal of Cancer Research, 83,
644–649.
116. Ariyoshi K, Hamada H, Naito M, et al. (1992) Mouse–
human chimeric antibody MH171 against the multi-
drug transporter P-glycoprotein. Japanese Journal of
117. Olsson L, Kaplan H. (1981) Human–human hybridoma
secretory monoclonal antibody on predefined anti-
gen specificity. Proceedings of the National Academy of
118. Isaccs JD, Watts RA, Hazleman BL, et al. (1992)
Humanized monoclonal antibody therapy for rheuma-
toid arthritis. Lancet, 340, 748–752.
119. Adair JR. (1992) Engineering antibodies for therapy.
Immunological Reviews, 130, 5–40.
120. Co MS, Avdalovic NM, Caron PC, Avdalovic MV,
Scheinberg DA, Queen C. (1992) Chimeric and
humanized antibodies with specificity for CD33 anti-
gen. Journal of Immunology, 148, 1149–1154.
121. Carter P, Presta L, Gorman CM, et al. (1992) Humani-
zation of an anti-P185HER2
antibody for human cancer
therapy. Proceedings of the National Academy of Sciences
of the USA, 89, 4285–4289.
122. Riechman L, Clark M, Waldmen H, Winter G. (1988)
Reshaping human antibodies for therapy. Nature, 332,
323–327.
123. Hale G, Dyer MJ, Clark MR, Phillips JM, Marcus R,
Reichmann L, Winter G, Waldmann N. (1988) Remis-
sion induction in non-Hodgkin’s lymphoma with
reshaped human monoclonal antibody CAMPATH-
1H. Lancet, ii, 1394–1399.
124. Mathieson PW, Cabbold SP, Hale G, et al. (1990)
Monoclonal antibody therapy in systemic vasculitis.
New England Journal of Medicine, 332, 250–254.
125. Tempest PR, Bnenner P, Lambert M, Taylor G, Furze
JM, Carr FJ, Harris WJ. (1991) Reshaping a human
monoclonal antibody to inhibit human respiratory
syncytial virus infection in vivo. Biological Technology,
9, 266–271.
126. Borrebaeck AK, Danielsson L, Moller SA. (1988)
Human monoclonal antibodies produced by primary in
vitro immunization of peripheral blood lymphocytes.
USA, 85, 3995–3999.

127. Neuberger MS, Williams GT, Fox RO. (1984) Recom-
binant antibodies possessing novel effector functions.
Nature, 312, 604–608.
128. Bode C, Matsueda G, Hui K, Haber E. (1985) Targeted
thrombolysis with a fibrin specific antibody–urokinase
conjugate. Science, 229, 765–767.
129. Schnee JM, Runge MS, Matsueda G, et al. (1987)
Construction and expression of a recombinant anti-
body targeted plasminogen activator. Proceedings of the
6908.
130. Byrn RA, Mordentii J, Lucas C, et al. (1990) Biological
properties of CD4 immunoadhesin. Nature, 344, 667–
670.
131. Capon DJ, Channow SM, Mordentii J, et al. (1989)
Designing CD4 immunoadhesin for AIDS therapy.
Nature, 337, 525–531.
132. Heike Y, Isuruo T. (1992) Augmentation by bispecific
F(ab*)2 reactive with P-glycoprotein and CD3 of
cytotoxicity of human effector cells in P-glycoprotein
positive human renal cancer cells. Japanese Journal of
133. Pearson CK, Cunningham C. (1993) Multidrug resist-
ance during cancer chemotherapy: biotechnological
solutions to a clinical problem. Trends in Biotechnology,
11, 511–516.
134. Jung G, Ledbetter JA, Muller-Eberhard HJ. (1987)
Induction of cytotoxicity in resting human
T-lymphocytes bound to tumor cells by antibody
heteroconjugates. Proceedings of the National Academic
of Sciences of the USA, 84, 4611–4615.
135. Drakemen DL, Fanger MW. (1995) Bispecific anti-
bodies and targeted cellular toxicity. Drug News and
Perspectives, 8, 189–192.
136. Fanger MW, Drakeman DL. (1995) Bispecific anti-
bodies. Drug News and Perspectives, 8, 133–137.
137. Ward ES, Gusson D, Griffith AD, Jones PT, Winter G.
(1989) Binding activities of a reportoire of single
immunoglobulin variable domain secreted from
Escherichia coli. Nature, 341, 544–546.
138. Brunt JV. (1990) Amplifying genes: PCR and its
alternatives. Biotechnology, 8, 291–294.
139. Griffiths AD, Malmqvist M, Marks JD, et al. (1993)
Human antidelf antibodies with high specificity from
phage display library. EMBO Journal, 12, 725–735.
140. Huse WD, Sastry L, Iverson SA, et al. (1989) Gener-
ation of a large combinatorial library of the immu-
noglobulins reportoire in phage lambda. Science, 246,
1275–1281.
141. Heaver GA. (1992) Monoclonal antibodies in
therapy. Annual Reports in Medicinal Chemistry, 27,
179–188.

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