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Tesi etd-08122014-114251

Thesis type
Tesi di laurea magistrale
Peptide-based targeted stealth liposomes: synthesis, in vitro evaluation, and application to contrast-enhanced ultrasonography
Corso di studi
relatore Prof. Beltram, Fabio
correlatore Dott. Signore, Giovanni
Parole chiave
  • peptide
  • stealth liposomes
  • liposomes
  • peptide aptamers
  • microbubbles
  • CEUS
  • contrast-enhanced ultrasound
Data inizio appello
Riassunto analitico
Efficiency of conventional antitumor therapies is often limited by unfavorable pharmacokinetics of conventional drugs, which usually show significant off-target activity, poor internalization in cells, and scarce retention in the host system. Nanomedicine aims at overcoming most of these limitations by allowing targeted, effective delivery of drugs and/or contrast agents, with the final goal of performing combined diagnostic and therapeutic activity with a single system, an approach referred to as theranostics; to this end, several nanostructures have been studied, but organic-based assemblies such as liposomes or lipoplexes are particularly attractive owing to their easy preparation, biocompatibility, and biodegradability.
Liposomes have been studied as drug carriers since the 1960s, owing to their ability to convey drugs to tumor tissues by passive accumulation, limiting side effects which are often encountered when dealing with free drugs. Notably, liposomal drug formulations are routinely used in a clinical setting. Efficiency of liposomal-based therapy can be significantly enhanced through insertion of poly-(ehtyleneglycol) residues, which inhibit protein adsorption and prevent opsonization, thus increasing residence time of the carrier in the bloodstream. An attractive improvement in liposome-based therapy is represented by the insertion of functional units that promote active targeting of a specific cell or tissue. Indeed, use of antibody-, ligand-, or peptide-derivatized liposomes represents a promising integration of already assessed delivery technologies with innovative targeting components.
In this work we developed a new class of liposomes tailored to in vivo targeted delivery and diagnostic. Our structures were assembled starting from commercially available lipids and novel lipopeptides, and their physic-chemical properties were evaluated in vitro and in living cells. We rationally designed and synthesized lipopeptides able to i) perform effective targeting of transferrin receptor, a membrane protein overexpressed by most tumor cells, and ii) hamper aspecific adsorption of serum proteins on the surface of the liposome, a key step in the recognition and clearance of nanostructures by the immune system.
Peptide-based liposomes were examined in detail, both in controlled conditions (cuvette) to assess their stability in various media, and in vitro on a model cell line of human pancreatic carcinoma (MIA PaCa-2) to investigate their biological activity; in both cases they compare favorably with plain, lipid-based liposomes already described in the literature.
Finally, the liposomes were conjugated to microbubbles, a contrast agent widely used in ultrasound based imaging, and their binding to MIA PaCa-2 cells was quantitated through flow cytometry.
We found that targeting properties of liposomes are fully retained upon conjugation to microbubbles, and thus they effectively recognize tumor cells. In perspective, our liposome-coated microbubbles can be regarded as a promising tool for in vivo theranostics.