earticle

영어

Development of agonistic monoclonal antibodies (mAbs) against the pro-apoptotic tumor necrosis factor (TNF)-related apoptosis inducing ligand (TRAIL) receptor 1, death receptor 4 (DR4), is an attractive anti-cancer strategy because of the tumorselective cell death-inducing activity. In this study, we first humanized the agonistic anti-DR4 AY4 scFv raised in mice (mAY4) by grafting the complementaritydetermining regions (CDRs) onto a fixed human framework, while preserving the so called Vernier zone residues, a group of framework (FR) residues directly underneath the CDRs, with the murine residues in the humanized antibody, hAY4. The humanized hAY4 scFv maintained the antigen binding affinity and specificity, including the antigen binding epitope, comparable to mAY4 scFv. To investigate how the valence of hAY4 scFv affect the DR4-mediated cell death, bivalent and trivalent
forms of hAY4 scFv were generated by linking with the dimerizing leucine zipper
and trimerizing isoleucine zipper coiled-coil domain via a hinge region, respectively.
Compared with the monovalent and bivalent forms, the trivalent hAY4 scFv induced
more potent caspase-dependent apoptotic cell death exhibiting the greater activations
of caspase-8 and the downstream pro-apoptotic molecules. Our results suggest that,
like other TNF family receptors, the avidity-mediated higher oligomerization of DR4
augment the receptor-mediated apoptotic cell death activity by promoting the intracellular cell death signaling.