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epitopes, which can be presented in the context of more than one HLA; such
epitopes are broadly recognized in human populations.
6.3.2 Decreasing Immunogenicity (Case Study)
Evidence that EpiMatrix can evaluate and identify epitopes that can be modified
to eliminate immunogenicity is provided by an analysis of SakSTAR and its
modified (deimmunized) derivative. SakSTAR is a natural variant of staphylo-
kinase, which is used as a potent thrombolytic agent in the setting of acute myo-
cardial infarction. Administration of SakSTAR results in the generation of IgG
antibodies, which limits the efficacy of the therapy. Warmerdam, Plaisance,
Vanderlick, Vandervoort, Brepoels, Collen, and De Maeyer (1998) identified the
“C3” region of SakSTAR (residues 71-87) as being highly reactive in patients
expressing either the DRB1*0301 or DRB1*0701 alleles. EpiMatrix parsed the
C3 region into overlapping 9-mer frames and scored each frame for MHC bind-
ing potential with respect to eight common alleles. Peptides contained between
residues 71 and 87 received the highest scores for the alleles (DRB1*0301 and
DRB1*0701) using the EpiMatrix algorithm, consistent with Warmerdam and
colleagues' observation that this region was highly immunogenic. The EpiMatrix
percentile ranking is shown in Table 3a.
Alanine substitutions to the MHC anchoring residues Y73, K74, R77, E80, and
D82, alone or in combination, were subsequently shown to reduce or eliminate T-cell
response. These modifications also resulted in dramatically lower EpiMatrix scores
for the protein, below the usual cutoff for immunogenicity (i.e., the 95
th
percentile).
An analysis of EpiMatrix scores for these peptides shows that Y73, K74, R77, E80,
and D82 all function as anchoring residues in our models of either DRB1*0701,
DRB1*0301, or both. Alanine substitutions at these positions had a neutral or nega-
tive effect on predicted binding affinity (EpiMatrix percentile ranking shown in
Table 3b).
In a separate study by Collen et al. (1996) the SakSTAR variant K74A, E75A,
R77A, E80A, D82A was shown to have “induced significantly fewer circulating
neutralizing antibodies” in human subjects. Again, this is consistent with the theory
that reduction of epitope content also reduces immunogenicity.
In summary, epitope mapping can uncover regions of therapeutic proteins in-
volved in the generation of antibody responses to such proteins. These regions can
Table 3a
. SakSTAR epitopes.
Original SakSTAR epitope
DRB1*0701 score: 99
th
percentile
73
YKEFRVVEL
81
DRB1*0301 score: 98
th
percentile
76
FRVVELDPS
84
DRB1*0301 score: 99
th
percentile
79
VELDPSAKI
87
