342
Antibody Molecules, Genetic Engineering of
4.2
Bispecifc Antibodies
In contrast to normal antibodies, which
are monospecifc, bispecifc antibodies
contain binding sites with two diFFerent
specifcities. Antibodies with two diFFer-
ent specifcities have been prepared by
chemical modifcation to combine univa-
lent Fragments oF diFFerent pepsin-treated
antibodies, by Fusing two hybridomas se-
creting antibodies oF diFFerent specifcities
yielding a quadroma and by joining two
diFFerent single-chain antibodies (sc±vs).
However, chemical modifcation oF an-
tibodies is ineFfcient and can lead to
side reactions that damage the combin-
ing site. In a quadroma, it is diFfcult to
separate the desired bispecifc antibodies
From the mixed population oF heavy and
light chains produced by the two hybrido-
mas, and sc±vs lacking constant regions
also lack the antibody eFFector Functions,
which may be critical in certain applica-
tions. To address these problems, novel
bispecifc antibodies have been produced
in which an sc±v oF one specifcity has
been genetically Fused aFter the hinge
(hinge-sc±v) or at the carboxy-terminus
(C
H
3-sc±v) oF an antibody with a diF-
Ferent specifcity. Both Fusion proteins
were expressed by gene transFection in
the context oF a murine variable region.
TransFectomas secreted a homogeneous
population oF the recombinant antibody
with the two diFFerent specifcities, one
at the amino-terminus (anti-dextran) and
the other at the carboxy-terminus (anti-
dansyl). The C
H
3-sc±v antibody, which
maintains the constant region oF human
IgG3, has some oF the associated eFFec-
tor Functions such as long halF-liFe and
±c receptor binding. As expected, hinge-
sc±v antibody, which lacks the C
H
2and
C
H
3 domains, has no known eFFector
Functions.
Bispecifc antibodies provide potential
tools For use in immunotherapy. They take
advantage oF the great specifcity oF vari-
able regions For their antigens and can
be envisioned as transporters oF therapeu-
tic drugs or molecules or even immune
eFFector cells to the specifc targets identi-
fed by one oF their binding sites. In Fact,
bispecifc antibodies have been shown to
be benefcial in the recruitment oF im-
mune cells For the treatment oF cancer.
Depending on the eFFector/target interac-
tion, bispecifc antibodies enhance cyto-
toxicity and phagocytosis. Several oF these
antibodies are currently undergoing eval-
uation in phase I and II clinical trials.
Among the most extensively studied bis-
pecifc antibodies are 2B1 and MDX-210.
2B1 produced From a hybrid hybridoma
is specifc For the tumor-associated anti-
gens HER2/
neu
and ±c
γ
RIII (CD16, the
±c gamma receptor expressed by key
cytotoxic eFFector cells such as natural
killer
cells,
neutrophils,
and
activated
mononuclear phagocytes), while MDX-
210 is a chemically conjugated hetero
±(ab)
0
2F
r
a
gm
en
tsp
e
c
if
cF
o
rHER
2
/
neu
and ±c
γ
RI (CD64, the ±c gamma receptor
expressed by key cytotoxic eFFector cells
such as monocytes, macrophages, and
I±N-
γ
–activated granulocytes). Both an-
tibodies showed therapeutic promise in
late-stage cancer patients whose tumor
was reFractory to conventional therapy.
Since conventionally prepared bispecifc
antibodies have already shown promise
in clinical trials and results From pre-
clinical studies oF recombinant bispecifc
antibodies are encouraging, in the Future
recombinant bispecifc antibodies will un-
doubtedly be used in multiple clinical
trials.
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