Molecular Engineering Strategies Tailoring the Apoptotic Response to a MET Therapeutic Antibody

Abstract

The MET oncogene encodes a tyrosine kinase receptor involved in the control of a complex network of biological responses that include protection from apoptosis and stimulation of cell growth during embryogenesis, tissue regeneration, and cancer progression. We previously developed an antagonist antibody (DN30) inducing the physical removal of the receptor from the cell surface and resulting in suppression of the biological responses to MET. In its bivalent form, the antibody displayed a residual agonist activity, due to dimerization of the lingering receptors, and partial activation of the downstream signaling cascade. The balance between the two opposing activities is variable in different biological systems and is hardly predictable. In this study, we generated and characterized two single-chain antibody fragments derived from DN30, sharing the same variable regions but including linkers different in length and composition. The two engineered molecules bind MET with high affinity but induce different biological responses. One behaves as a MET-antagonist, promoting programmed cell death in MET "addicted" cancer cells. The other acts as a hepatocyte growth factor (HGF)-mimetic, protecting normal cells from doxorubicin-induced apoptosis. Thus, by engineering the same receptor antibody, it is possible to generate molecules enhancing or inhibiting apoptosis either to kill cancer cells or to protect healthy tissues from the injuries of chemotherapy.

A receptor-antibody hybrid hampering MET-driven metastatic spread

“…Recombinant fusion proteins were designed starting from the sequence of MvDN30_long-sc60 [20] and decoyMET K842E [17], joining the two moieties by an amino acid sequence (linker). DecAb fusion proteins were obtained connecting the IPT4 domain of decoy-MET K842E to the variable domain of the light chain of MvDN30_long-sc60.…”

“…To generate a recombinant fusion that is efficiently produced and assembled, the antibody must be in single chain format. We previously designed and validated MvDN30_long-sc60, a single chain Fab derived from DN30 that maintains MET inhibitory properties of the precursor antibody [20]. To prevent the reciprocal neutralization of the moieties by intra-chain binding, the MET epitope bound by DN30, the lysine residue at position 842 [17], was converted into a glutamic acid (decoyMET K842E ).…”