92
Combinatorial Phage Antibody Libraries
of
β
-sheet content. Elicitation of antibodies
against prion protein is difFcult due to host
tolerance of endogenous PrP
C
.Thisprob
-
lem was overcome by immunizing mice in
which the
PrP
gene was ablated (Prnp
0
/
0
)
,
and antibodies were retrieved from phage
display libraries prepared from the bone
marrow and spleens of these mice. ±abs
speciFc for both discontinuous and lin-
ear epitopes covering the majority of the
murine prion molecule were isolated, most
of which were cross-reactive with human
PrP. Surprisingly, although the mice were
immunized with infectious PrP fragment
preparations, ±abs against only the cellu-
lar form of PrP were isolated (±ig. 7). The
antibodies generated have given insight
into the conformational changes associ-
ated with PrP. ±or example, several of the
antibodies were used to show that PrP
that is sparsely immobilized on a biosen-
sor chip surface can undergo spontaneous
rearrangement to a conformation similar
to PrP
Sc
. More importantly, one of the
±abs, D18, has been shown to bind tightly
to cell-surface PrP
C
and inhibit PrP
Sc
formation in a dose-dependent manner.
These results may eventually lead to the
prevention or treatment of prion diseases
by anti-PrP antibodies.
4.1.2
Antibodies against
Tumor-associated Antigens
Combinatorial antibody libraries are be-
ing increasingly used in cancer research
in which an increasing number of tumor-
associated antigens are being identiFed
as
potential
targets
for
disease
inter-
vention. The successes in treatment of
certain cancers with monoclonal antibod-
ies such as in the treatment of relapsed
HER2 breast malignancy by Herceptin
(Genentech), and B-cell follicular lym-
phoma by Rituximab (IDEC/Genentech),
have set a precedence that could be fol-
lowed by library-derived antibodies in the
near future. There are many examples of
phage display–selected antibodies against
tumor-associated antigens, but we will
mention only a few due to limitation of
space. Of particular note are studies in
which phage display–selected antibodies
E123
E149
D13
R1,R2
D18, 28d
Fig. 7
Regions of prion protein (PrP)
bound by individual antibodies selected
from phage display libraries. Each antibody
epitope has been assigned with a rectangle
of different color as indicated on the Fgure.
Beginning at the N-terminus, E123: purple;
E149: grey; D13: blue; 3±4: yellow; 13A5:
black line; D18 and 28d: red; R1 and R2:
green. The cell membrane is shown as a
black line at the bottom of the Fgure. The
glycosylated side chains attached to Asn
181 and Asn197 are shown in orange and
yellow respectively. The glycosyl
phosphatidyl inositol protein (GPI) group
anchoring PrP to the cell surface is shown
in light blue. The antibody D18 has been
shown to inhibit the formation of the
infectious PrP
Sc
form of the protein in a
dose-dependent manner. [Adapted from
Leclerc et al. (2003)
J. Mol. Biol.
326
,
475–83.] (See color plate p. xxv.)
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