Drug Conjugation Induced Modulation of Structural and Membrane Interaction Features of Cationic Cell-Permeable Peptides

Abstract

Cell-penetrating peptides might have great potential for enhancing the therapeutic effect of drug molecules against such dangerous pathogens as Mycobacterium tuberculosis (Mtb), which causes a major health problem worldwide. A set of cationic cell-penetration peptides with various hydrophobicity were selected and synthesized as drug carrier of isoniazid (INH), a first-line antibacterial agent against tuberculosis. Molecular interactions between the peptides and their INH-conjugates with cell-membrane-forming lipid layers composed of DPPC and mycolic acid (a characteristic component of Mtb cell wall) were evaluated, using the Langmuir balance technique. Secondary structure of the INH conjugates was analyzed and compared to that of the native peptides by circular dichroism spectroscopic experiments performed in aqueous and membrane mimetic environment. A correlation was found between the conjugation induced conformational and membrane affinity changes of the INH–peptide conjugates. The degree and mode of interaction were also characterized by AFM imaging of penetrated lipid layers. In vitro biological evaluation was performed with Penetratin and Transportan conjugates. Results showed similar internalization rate into EBC-1 human squamous cell carcinoma, but markedly different subcellular localization and activity on intracellular Mtb.

Cellular Internalization and Inhibition Capacity of New Anti-Glioma Peptide Conjugates: Physicochemical Characterization and Evaluation on Various Monolayer- and 3D-Spheroid-Based in Vitro Platforms

“…The higher membrane affinity of the SynB3-containing Dau-conjugates observed in lipid monolayer experiments is in harmony with their conformational change in membrane mimetic solvent (i.e., increased α-helix content) determined by CD measurements (Table ). A similar correlation was found to be valid previously for structural and membrane interaction features of cationic CPPs …”

“…A similar correlation was found to be valid previously for structural and membrane interaction features of cationic CPPs. 95 We studied the penetration ability of the conjugates on Transwell (TW) using co-cultured noncontact monolayers of HUVEC and U87 cells as a simple in vitro BBB model (Figure 10C,D based on ref 96). Compounds (Dau and conjugates Dau-Aoa-SynB3, Dau-Aoa-TKPPR, and Dau-Aoa-SynB3-TKPPR) were added to the HUVEC-containing apical chamber at 50 μM concentration, and after 45 min or 3 h of incubation, U87 cells from the basolateral chamber were stained and fixed prior to CLSM analysis.…”

Novel Assay Platform to Evaluate Intracellular Killing of Mycobacterium tuberculosis: In Vitro and In Vivo Validation

“…These features strongly influence the internalization and intracellular activity of the compounds. The interaction of INH-peptide conjugates with lipid monolayers was the topic of our previous work ( 70 ). As a result, we could conclude that the degree of penetration into a TB membrane model, consisting of DPPC+mycolic acid (3:1), was the highest for INH-transportan, INH-CM15, INH-magainin, and INH-Dhvar4 conjugates.…”

Cell-Penetrating Peptides and Transportan

“…A few reports are available on the characterization of the interactions of transportan with different lipid membranes. CD spectroscopy showed that the secondary structure of transportan and penetratin, isoniazid–penetratin and –transportan showed increased membrane affinity with DPPC and DPPC + mycolic acid mixed monolayers [ 126 ]. The binding of transportan 10 and its four variants to phospholipid vesicles showed the peptide-induced efflux, “becoming faster as the Gibbs energies for binding and insertion of the TP10 variants decrease” [ 134 ].…”