Aib-containing peptide analogs: cellular uptake and utilization in oligonucleotide delivery

Area of Science:

• Biochemistry
• Molecular Biology
• Peptide Chemistry

Background:

• Cell-penetrating peptides (CPPs) facilitate cellular uptake of molecules.
• Structure-activity relationships of CPPs are crucial for optimizing delivery.
• α-Aminoisobutyric acid (Aib) can modulate peptide structure and function.

Purpose of the Study:

• To synthesize and evaluate α-Aminoisobutyric acid (Aib)-containing peptide analogs of TV-XIIa for enhanced cellular uptake.
• To investigate the potential of these peptide analogs for delivering oligonucleotides (ODNs).
• To assess the antisense effect of a peptide-oligonucleotide conjugate on gene expression.

Main Methods:

• Synthesis of peptide analogs by substituting specific amino acids (Aib, Ala, Pro) in the TV-XIIa sequence.
• Creation of a covalent conjugate (P-IV-AON) between a modified peptide (P-IV) and a 15-mer antisense ODN.
• Evaluation of cellular uptake and antisense activity in A549 cells using luciferase gene expression assays.

Main Results:

• Substitution of Aib at positions 1, 5, or 9 with alanine (Ala) reduced cellular uptake.
• Simultaneous substitution of proline (Pro) residues at positions 6 and 10 with Aib (P-IV) significantly increased cellular uptake.
• The P-IV-AON conjugate demonstrated a dose-dependent decrease in luciferase expression at 5 and 10 μM concentrations.

Conclusions:

• α-Aminoisobutyric acid incorporation can significantly enhance the cell-penetrating properties of peptide analogs.
• The P-IV peptide analog shows promise for developing effective oligonucleotide delivery systems.
• The P-IV-AON conjugate exhibits functional antisense activity, indicating its potential for therapeutic applications.