A Helix-Stabilizing Linker Improves Subcutaneous Bioavailability of a Helical Peptide Independent of Linker Lipophilicity

Abstract

Stabilized peptides address several limitations to peptide-based imaging agents and therapeutics such as poor stability and low affinity due to conformational flexibility. There is also active research in developing these compounds for intracellular drug targeting, and significant efforts have been invested to determine the effects of helix stabilization on intracellular delivery. However, much less is known about the impact on other pharmacokinetic parameters such as plasma clearance and bioavailability. We investigated the effect of different fluorescent helix-stabilizing linkers with varying lipophilicity on subcutaneous (SC) bioavailability using the glucagon-like peptide-1 (GLP-1) receptor ligand exendin as a model system. The stabilized peptides showed significantly higher protease resistance and increased bioavailability independent of linker hydrophilicity, and all subcutaneously delivered conjugates were able to successfully target the islets of Langerhans with high specificity. The lipophilic peptide variants had slower absorption and plasma clearance than their respective hydrophilic conjugates, and the absolute bioavailability was also lower likely due to the longer residence times in the skin. The ease and efficiency of double-click helix stabilization chemistries is a useful tool for increasing the bioavailability of peptide therapeutics, many of which suffer from rapid in vivo protease degradation. Helix stabilization using linkers of varying lipophilicity can further control SC absorption and clearance rates to customize plasma pharmacokinetics.

Stapled ghrelin peptides as fluorescent imaging probes

“…This was unexpected, as the addition of Cy5 to the lead i, i + 4 and i, i + 7 sequences ( 24 and 25 , respectively), did not elicit the same increase in helicity (Figure ). It has been reported that cyanine dyes can increase the helicity of peptide sequences, possibly due to the lipophilic structure of the dye interacting with side chains . It is also worth noting that compounds 23–25 do not have the characteristic maxima below 200 nm that is expected of an α‐helix.…”

Blocking of Glucagonlike Peptide-1 Receptors in the Exocrine Pancreas Improves Specificity for β-Cells in a Mouse Model of Type 1 Diabetes

“…All chemicals were purchased from Sigma Aldrich, unless otherwise specified. Peptide and reaction details are provided in Supplemental Figure 1 (supplemental materials are available at http://jnm.snmjournals.org) (16,22). All animal experiments were conducted under the approval of the Institutional Animal Care and Use Committee at the University of Michigan and Memorial Sloan Kettering Cancer Center and followed the National Institutes of Health guidelines for animal welfare.…”

Tracking Antibody Distribution with Near-Infrared Fluorescent Dyes: Impact of Dye Structure and Degree of Labeling on Plasma Clearance

“…After purification, conjugates were run on SDS-PAGE and scanned on an Odyssey CLx to ensure all free dye was removed (Figure S2). Binding affinities were performed as previously described 3 using HCC1954 cells. Briefly, titrations of unlabeled antibody and antibody–dye conjugates were incubated with 50,000 HCC1954 cells on ice for 3 h and washed.…”