Controlled Release of rAAV Vectors from APMA-Functionalized Contact Lenses for Corneal Gene Therapy.

Fernando Alvarez-Rivera,Angel Concheiro,Carmen Alvarez-Lorenzo,Jagadeesh K Venkatesan,Ana Rey-Rico,Magali Cucchiarini,Luis Diaz-Gomez

doi:10.3390/pharmaceutics12040335

Fernando Alvarez-Rivera, Angel Concheiro + Show 5 more

Open Access

https://doi.org/10.3390/pharmaceutics12040335

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Abstract

As an alternative to eye drops and ocular injections for gene therapy, the aim of this work was to design for the first time hydrogel contact lenses that can act as platforms for the controlled delivery of viral vectors (recombinant adeno-associated virus, rAAV) to the eye in an effective way with improved patient compliance. Hydrogels of hydroxyethyl methacrylate (HEMA) with aminopropyl methacrylamide (APMA) (H1: 40, and H2: 80 mM) or without (Hc: 0 mM) were synthesized, sterilized by steam heat (121 °C, 20 min), and then tested for gene therapy using rAAV vectors to deliver the genes to the cornea. The hydrogels showed adequate light transparency, oxygen permeability, and swelling for use as contact lenses. Loading of viral vectors (rAAV-lacZ, rAAV-RFP, or rAAV-hIGF-I) was carried out at 4 °C to maintain viral vector titer. Release in culture medium was monitored by fluorescence with Cy3-rAAV-lacZ and AAV Titration ELISA. Transduction efficacy was tested through reporter genes lacZ and RFP in human bone marrow derived mesenchymal stem cells (hMSCs). lacZ was detected with X-Gal staining and quantified with Beta-Glo®, and RFP was monitored by fluorescence. The ability of rAAV-hIGF-I-loaded hydrogels to trigger cell proliferation in hMSCs was evaluated by immunohistochemistry. Finally, the ability of rAAV-lacZ-loaded hydrogels to transduce bovine cornea was confirmed through detection with X-Gal staining of β-galactosidase expressed within the tissue.

Highlights

Ocular gene therapy has the potential to cure or relieve symptoms of inherited or acquired diseases by replacing a defective gene with a normal gene [1,2]
The aim of this work was to provide a proof-of-principle of the suitability of functionalized poly(2-hydroxyethyl methacrylate) (p-HEMA) hydrogels as platforms that host and release recombinant adeno-associated viral vectors (rAAV) vectors for sustained gene therapy of corneal complications, while keeping adequate mechanical and optical properties to act as ocular bandages or Contact lenses (CLs)
Swelling behavior of autoclaved discs was similar after 24 h storage at 4 ◦C; hydrogel discs (Hc): 52.0 (s.d. 1.5)%; H1: 49.7 (s.d. 1.4)%; and H2: 51.6 (s.d. 0.8)% (Figure 2a,b)

Summary

Introduction

Ocular gene therapy has the potential to cure or relieve symptoms of inherited or acquired diseases by replacing a defective gene with a normal gene [1,2] This chance to cure by correcting the cause of the disorder means an important advantage compared with conventional drugs that, in most cases, only can suppress symptoms [3,4]. Cornea is an avascular tissue that refracts the light (together with the lens) and acts as a first protective barrier due to the tight junctions of the epithelium and Bowman’s membrane This important barrier function along with other eye protective mechanisms (e.g., lachrymal fluid replacement, blinking, nasolachrymal drainage, etc.) represents a challenge to achieve sufficient drug bioavailability in the ocular structures after topical application of eye drops [9,11]

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