PK modulation of haptenylated peptides via non-covalent antibody complexation

Abstract

We applied noncovalent complexes of digoxigenin (Dig) binding antibodies with digoxigeninylated peptide derivatives to modulate their pharmacokinetic properties. A peptide derivative which activates the Y2R receptor was selectively mono-digoxigeninylated by reacting a NHS-Dig derivative with an ε-amino group of lysine 2. This position tolerates modifications without destroying receptor binding and functionality of the peptide. Dig-peptide derivatives can be loaded onto Dig-binding IgGs in a simple and robust reaction, thereby generating peptide-IgG complexes in a defined two to one molar ratio. This indicates that each antibody arm becomes occupied by one haptenylated peptide. In vitro receptor binding and signaling assays showed that Dig-peptides as well as the peptide-antibody complexes retain better potency than the corresponding pegylated peptides. In vivo analyses revealed prolonged serum half-life of antibody-complexed peptides compared to unmodified peptides. Thus, complexes are of sufficient stability for PK modulation. We observed more prolonged weight reduction in a murine diet-induced obesity (DIO) model with antibody-complexed peptides compared to unmodified peptides. We conclude that antibody-hapten complexation can be applied to modulate the PK of haptenylated peptides and in consequence improve the therapeutic efficacy of therapeutic peptides.

Hapten‐directed spontaneous disulfide shuffling: a universal technology for site‐directed covalent coupling of payloads to antibodies

“…One objective to covalently link payloads to hapten binding antibodies is to increase the stability between payload and antibody in vivo . We have previously shown that noncovalent connections between antibodies and haptenylated payloads can be applied to prolong the serum half-life of small compounds and peptides ( 17 ). In these experiments, we observed a profound increase of the serum half-life of the payload, and we also observed release of the noncovalently linked payload from the antibody after extended periods of time.…”

“…Final cleavage was performed as described above. The crude product was purified by preparative reversed phase HPLC as previously described ( 17 ) giving a colorless solid ( m / z : calculated for C131H197N39O30S2 = 2862.4; found: 2862.4).…”

“…We have recently devised another approach to couple small compounds and peptides to antibodies and thereby modulate their pharmacokinetics. This technology bases upon haptenylated payloads that are complexes by antibodies in a noncovalent manner ( 17 ). In contrast to stable covalent conjugates, noncovalent binding (whose strength is determined by on and off rates of the hapten binder) enables payload release as well as recapture in the circulation.…”

“…Although the noncovalent attachment is suitable to significantly prolong the serum half-life, liberation will over time lead to payload elimination because not all payload rebinds to the antibody ( 17 ). To address this issue and devise a hapten-based platform that has payloads even more stably attached to antibodies, we designed a disulfide bond between antibody and payload to stabilize the antibody-hapten-payload complex.…”

Engineered hapten‐binding antibody derivatives for modulation of pharmacokinetic properties of small molecules and targeted payload delivery

“…These animals were treated with different NPY2 receptor agonists, known to be able to modulate the obese phenotype (provided active concentrations can be achieved for sufficient time) . Hoffmann and coworkers could demonstrate that the peptide PYY or Dig‐coupled PYY derivatives by itself had very limited potency (due to rapid elimination), but had improved potency when it was complexed with an anti‐digoxigenin antibody . Obese mice treated with the PYY‐Dig–antibody complexes showed reduced food intake even after 48 h, whereas peptide alone showed efficacy only in the first hour after treatment .…”