Auristatins are highly potent
antimitotic agents that have received considerable attention because of their activities when targeted to
tumor cells in the form of
antibody-drug conjugates (ADCs). Our lead agent,
SGN-35, consists of the cAC10 antibody linked to the N-terminal
amino acid of
monomethylauristatin E (MMAE) via a
valine-
citrulline p-aminobenzylcarbamate (val-
cit-
PABC) linker that is cleaved by intracellular
proteases such as
cathepsin B. More recently, we developed an
auristatin F (AF) derivative
monomethylauristatin F (MMAF), which unlike MMAE contains the
amino acid phenylalanine at the C-terminal position. Because of the negatively charged C-terminal residue, the potency of AF and MMAF is impaired. However, their ability to kill target cells is greatly enhanced through facilitated cellular uptake by internalizing mAbs. Here, we explore the effects of linker technology on AF-based ADC potency, activity, and tolerability by generating a diverse set of
dipeptide linkers between the C-terminal residue and the mAb carrier. The resulting ADCs differed widely in activity, with some having significantly improved therapeutic indices compared to the original mAb-Val-
Cit-
PABC-MMAF conjugate. The therapeutic index was increased yet further by generating
dipeptide-based ADCs utilizing new auristatins with
methionine or
tryptophan as the C-terminal drug residue. These results demonstrate that manipulation of the C-terminal
peptide sequence used to attach auristatins to the mAb carrier can lead to highly potent and specific conjugates with greatly improved therapeutic windows.