Evaluation of GHK peptide–heparin interactions in multifunctional liposomal covering

Abstract

Small biospecific peptides with defined chemical structure and cellular responses are promising alternatives to full-length therapeutic proteins. Identification of these peptides solely or in combination with other bioactive factors and determination of their targets are of substantial interest in current drug delivery research. This study is aimed at the development of new liposomal formulations of ECM-derived GHK peptide known for its multiple regeneration-related activities but poorly recognized cellular targets. In situ association of membranotropic GHK derivative with unilamellar liposomes was performed to prepare GHK-modified liposomes with defined properties. According to DLS, the GHK component on the liposomal surface interacted with heparin in a specific manner compared to other polysaccharides and RGD counterpart, whereas ITC analysis of such interactions was complicated. The results provide a useful tool for screening of bio-interactions of synthetic peptide-presenting liposomes by the DLS technique. They were also employed to produce a multi-functional nanosized GHK-heparin covering for liposomes. The resulting composite liposomes possessed low size dispersity, increased anionic charge, and mechanical rigidity. The heparin component significantly promoted the accumulation of GHK-modified liposomes in 3T3 fibroblasts so that the composite liposomes exhibited the highest cell-penetrating activity. Furthermore, the latter formulation stimulated cell proliferation and strongly inhibited ROS production and GSH depletion under oxidative stress conditions. Together, the results support that cell-surface glycosaminoglycans can be involved in GHK-mediated liposomal delivery, which can be further greatly enhanced by association with heparin. The composite liposomes with GHK-heparin covering can be considered as an advanced GHK-based formulation for therapeutic and cosmeceutical applications.

Keywords:

• Peptide-presenting liposomes
• GHK-heparin interaction
• multi-functional liposomal covering
• targeted delivery
• cell stimulation and protection

Acknowledgement

The authors thank Vladimir G. Evtugyn, Elvira T. Siraeva, and Amina G. Daminova (Interdisciplinary Center for Analytical Microscopy of Kazan Federal University) for microscopy analysis. This work is part of the Kazan Federal University strategic academic leadership program.

Disclosure statement

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Data availability statement

The data presented in the study are available within the article and Supplementary Material.

Additional information

Funding

The study was funded by the Russian Science Foundation according to the research (project no. 20-73-10105) (preparation of peptide and liposome formulations and study of their interactions with polysaccharides and the cells). The study was supported by the Subsidy Allocated to Kazan Federal University for the State Assignment in the Sphere of Scientific Activities (project FZSM-2022–0020) (evaluation of primary cellular effects of the formulations).