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Blood, Vol. 95 No. 2 (January 15), 2000:
pp. 564-568
Reduction of the antigenicity of factor VIII toward complex
inhibitory antibody plasmas using multiply-substituted hybrid
human/porcine factor VIII molecules
Rachel T. Barrow,
John F. Healey,
David Gailani,
Dorothea Scandella, and
Pete Lollar
From Emory University, Atlanta, GA; the Division of
Hematology, Vanderbilt University, Nashville, TN; and the Holland
Laboratory, American Red Cross, Rockville, MD.
Factor VIII (fVIII) circulates as a heavy chain/light chain
(A1-A2-B/ap-A3-C1-C2) heterodimer. The 41-residue light chain activation peptide, ap, is cleaved from fVIII during
proteolytic activation by thrombin or factor Xa. We constructed 7 active recombinant hybrid B-domainless human/porcine fVIII molecules
that contained combinations of porcine sequence replacements within the
A2, ap-A3, and C2 domains. The cross-reactivity of 23 high-titer inhibitory antibodies between human fVIII and the hybrids
was inversely related to the degree of porcine substitution. In all
plasmas, the substitution of all 3 regions yielded cross-reactivities
that were not significantly different from those of porcine fVIII. To
differentiate between inhibitor binding to the ap region and
the A3 domain, we constructed 2 additional hybrids that contained
porcine A2 and C2 domain substitutions and either porcine A3 or porcine
ap substitutions. The porcine ap segment was less
antigenic than the human ap segment in several plasmas that had
activity against the ap-A3 region. This indicates that some
inhibitor plasmas contain antibodies directed against the fVIII
ap segment in addition to A2, A3, and C2 domain epitopes identified in previous studies. Substitution of porcine sequences within the A2, A3, C2, and ap regions of human fVIII is
necessary and sufficient to achieve a maximal reduction in antigenicity relative to porcine fVIII with respect to most inhibitory antibody plasmas.
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