Antibody–Drug Conjugate that Exhibits Synergistic Cytotoxicity with an Endosome–Disruptive Peptide

Antibody–drug
conjugates are an important class of cancer
therapeutics. These agents generally bind a specific cell surface
receptor, undergo receptor-mediated endocytosis, and enter the endosomal–lysosomal
system, where the environment in these organelles facilitates the
release of a membrane-permeable cytotoxin. By using a membrane-impermeable
cytotoxin, we describe here a method that allows the cytotoxicity
of an antibody conjugate to be triggered by co-administration with
an endosome-disruptive peptide that exhibits low toxicity. This approach
was validated by conjugation of an anionic derivative of the tubulin-binding
cytotoxin colchinol methyl ether to lysine residues of the HER2-targeting
antibody trastuzumab (Herceptin) via a disulfide. When this antibody
binds HER2 on SKBR3 breast cancer cells and undergoes endocytosis,
the membrane-impermeable cytotoxin is released, but it becomes trapped
in endosomes, resulting in relatively low cytotoxicity (IC50 > 1 μM). However, co-administration with an essentially
nontoxic
(IC50 > 10 μM) cholesterol-linked endosome-disruptive
peptide promotes the release of this small molecule into the cytoplasm,
conferring subnanomolar cytotoxic potency (IC50 = 0.11
± 0.07 nM). Studies of a structurally related fluorophore conjugate
revealed that the endosome-disruptive peptide does not substantially
enhance cleavage of the disulfide (t1/2 = 8 ± 2 h) within endosomes, suggesting that the mechanism
of endosomal escape involves the efflux of some small molecules without
facilitating substantial influx of reduced glutathione.