Immunoliposomes for Targeted Delivery of an Antifibrotic Drug

Abstract

Excessive extracellular matrix formation in organs and tissues arises from an imbalance between the synthesis and degradation of matrix proteins, especially collagen. This condition interferes with proper wound healing and regeneration, and to date, no specific treatment is available. In the present study, we propose a targeted drug delivery system consisting of cell-specific immunoliposomes (ILs) loaded with deferoxamine (DFO) as an antifibrotic drug. ILs were functionalized with polyethylene glycol (PEG) to improve the steric stability and prolong their half-life. In addition, a single-chain Fv (scFv) antibody fragment that specifically targets fibroblast activation protein (FAP) was incorporated. An in vitro fibrosis model was employed to test this construct. This model consisted of highly activated pro-fibrotic fibroblasts with 2- to 6-fold induction of selected fibrosis markers: cell/matrix deposited collagen I, total soluble collagen, and α smooth muscle actin. The activation was accompanied by a significant and cell-specific elevation of FAP expression and activity, thereby confirming that FAP is an adequate target for antifibrotic drug delivery. Purified anti-FAP scFv was shown to bind specifically to these cells without influencing the FAP enzymatic activity. DFO was demonstrated to have a dose-dependent antifibrotic activity as quantified by collagen deposition. Specific binding and intracellular uptake of DiI-labeled ILs into the activated fibroblasts were shown by flow cytometry and microscopy. Finally, DFO-loaded ILs targeted to FAP caused a significant reduction in the collagen deposition, whereas no effect was observed using liposomes that lacked the targeting antibody fragment. These results suggest that the FAP-specific scFv-conjugated liposomes have considerable potential for cell-specific targeting applicable as a therapy for excessive collagen deposition during fibrosis. In general, through liposome encapsulation, bioactive molecules, such as DFO, that have broad effects and poor cell penetration can be converted into cell-specific composites for targeted drug delivery.

Use of Single-Chain Antibody Derivatives for Targeted Drug Delivery

“…Unlike deferoxamine-loaded nontargeted liposomes, which were not able to decrease collagen deposition, deferoxamine-loaded ILs significantly reduced collagen deposition ( Figure 3C). Free deferoxamine did not reduce collagen deposition (53). These results indicate that anti-FAP scFv plays a significant role in delivering deferoxamine molecules to fibroblasts.…”

“…Fibroblasts of normal adult tissues express undetectable levels of FAP (55). To generate an in vitro model of fibrosis, Schuster and colleagues isolated primary normal human lung fibroblasts from adult lung tissue and treated them with transforming growth factor β1 (TGF β1; to induce fibrosis-stimulating conditions), and ascorbate and proline (for collagen synthesis) (53). Unlike deferoxamine-loaded nontargeted liposomes, which were not able to decrease collagen deposition, deferoxamine-loaded ILs significantly reduced collagen deposition ( Figure 3C).…”

“…To overcome these problems, Schuster and colleagues encapsulated deferoxamine molecules in PEG-functionalized ILs (scFv-conjugated liposomes) that were able to specifically target FAP-expressing cells. The scFv contained an additional cysteine residue within its linker region that allowed site-directed conjugation to liposomes (53). A cysteine residue within linker region serves as a suitable site for conjugation ( Figure 3C).…”

Evolution of Nanotechnology in Delivering Drugs to Eyes, Skin and Wounds via Topical Route

“…Conventional liposomes generally have an outer lipid layer and an inner aqueous core, whereas the new generation liposomes-also known as stealth liposomes-are coated with polyethylene glycol (PEG), and protect them from engulfment by phagocytic cells [27]. Another class of liposomes are immunoliposomes, which have been developed for targeted delivery by conjugating antibodies at the surface that recognizes specific proteins on the target cells [28][29][30]. Cationic liposomes consist of positively charged lipids that interact efficiently with the negatively charged DNA and thereby condense the DNA into a more compact structure; hence, they are mostly used for delivering genes into eyes into various diseases such as glaucoma, epithelial retinal diseases, cytomegalovirus retinitis, etc.…”

Liposome-targeted delivery for highly potent drugs

“…Thus, an efficient delivery system that intensifies its cell-specific bioavailability is essential. Schuster et al have developed DFO-loaded PEGylated immunoliposomes that specifically targets fibroblast activation protein (FAP) by single-chain fragment variable (scFv) (Schuster et al 2015). It has exhibited specific binding and uptake into FAP-expressing cells and significant reduction in the collagen deposition.…”