Novel Small Multilamellar Liposomes Containing Large Quantities of Peptide Nucleic Acid Selectively Kill Breast Cancer Cells

Galina Proshkina, Elena Shramova, Anastasiya Ryabova, Liat Katrivas, Clelia Giannini*, Daniele Malpicci, Yael Levi-Kalisman, Sergey Deyev, Alexander Kotlyar*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

1 Scopus citations


Peptide nucleic acid (PNA) may be used in various biomedical applications; however, these are currently limited, due to its low solubility in aqueous solutions. In this study, a methodology to overcome this limitation is demonstrated, as well as the effect of PNA on cell viability. We show that extruding a mixture of natural phospholipids and short (6–22 bases), cytosine-rich PNA through a 100 nm pore size membrane under mild acidic conditions resulted in the formation of small (60–90 nm in diameter) multilamellar vesicles (SMVs) comprising several (3–5) concentric lipid membranes. The PNA molecules, being positively charged under acidic conditions (due to protonation of cytosine bases in the sequence), bind electrostatically to negatively charged phospholipid membranes. The large membrane surface area allowed the encapsulation of thousands of PNA molecules in the vesicle. SMVs were conjugated with the designed ankyrin repeat protein (DARPin_9-29), which interacts with human epidermal growth factor receptor 2 (HER2), overexpressed in human breast cancer. The conjugate was shown to enter HER2-overexpressing cells by receptor-mediated endocytosis. PNA molecules, released from lysosomes, aggregate in the cytoplasm into micron-sized particles, which interfere with normal cell functioning, causing cell death. The ability of DARPin-functionalized SMVs to specifically deliver large quantities of PNA to cancer cells opens a new promising avenue for cancer therapy.

Original languageAmerican English
Article number4806
Issue number19
StatePublished - Oct 2022

Bibliographical note

Funding Information:
The authors acknowledge support by the Russian Science Foundation (grant no. 19-14-00112 in part of in vitro experiments), Russian Foundation for Basic Research (grant no. 20-04-00489 in part of flow cytometry experiments), Russian Science Foundation (grant no.21-74-30016 in part of confocal microscopy), and the Ministry of Science and Technology of Israel (grant no. 3-16495).

Publisher Copyright:
© 2022 by the authors.


  • DARPins
  • HER2
  • liposomes
  • peptide nucleic acid
  • targeting delivery


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