PLoS ONE 8(6): e67091. doi:10.1371/journal.pone.0067091
Development of Adenoviral Delivery Systems to Target Hepatic Stellate Cells In Vivo.
Reetz J, Genz B, Meier C, Kowtharapu BS, Timm F, Vollmar B, Herchenröder O, Abshagen K, Pützer BM.
Hepatic stellate cells (HSCs) are known as initiator cells that induce liver fibrosis upon intoxication or other noxes.
Deactivation of this ongoing remodeling process of liver parenchyma into fibrotic tissue induced by HSCs is an interesting
goal to be achieved by targeted genetic modification of HSCs. The most widely applied approach in gene therapy is the
utilization of specifically targeted vectors based on Adenovirus (Ad) serotype 5. To narrow down the otherwise ubiquitous
tropism of parental Ad, two modifications are required: a) ablating the native tropism and b) redirecting the vector particles
towards a specific entity solely present on the cells of interest. Therefore, we designed a peptide of the nerve growth factor
(NGFp) with specific affinity for the p75 neurotrophin receptor (p75NTR) present on HSCs. Coupling of this NGFp to vector
particles was done either via chemical conjugation using bifunctional polyethylene glycol (PEG) or, alternatively, by
molecular bridging with a fusion protein specific for viral fiber knob and p75NTR. Both Ad vectors transmit the gene for the
green fluorescent protein (GFP). GFP expression was monitored in vitro on primary murine HSCs as well as after systemic
administration in mice with healthy and fibrotic livers using intravital fluorescence microscopy. Coupling of NGFp to Ad via
S11 and/or PEGylation resulted in markedly reduced liver tropism and an enhanced adenoviral-mediated gene transfer to
HSCs. Transduction efficiency of both specific Ads was uniformly higher in fibrotic livers, whereas Ad.GFP-S11-NGFp
transduce activated HSCs better than Ad.GFP-PEG-NGFp. These experiments contribute to the development of a targeted
gene transfer system to specifically deliver antifibrotic compounds into activated HSCs by systemically applied adenoviral
vector modified with NGFp.
Deactivation of this ongoing remodeling process of liver parenchyma into fibrotic tissue induced by HSCs is an interesting
goal to be achieved by targeted genetic modification of HSCs. The most widely applied approach in gene therapy is the
utilization of specifically targeted vectors based on Adenovirus (Ad) serotype 5. To narrow down the otherwise ubiquitous
tropism of parental Ad, two modifications are required: a) ablating the native tropism and b) redirecting the vector particles
towards a specific entity solely present on the cells of interest. Therefore, we designed a peptide of the nerve growth factor
(NGFp) with specific affinity for the p75 neurotrophin receptor (p75NTR) present on HSCs. Coupling of this NGFp to vector
particles was done either via chemical conjugation using bifunctional polyethylene glycol (PEG) or, alternatively, by
molecular bridging with a fusion protein specific for viral fiber knob and p75NTR. Both Ad vectors transmit the gene for the
green fluorescent protein (GFP). GFP expression was monitored in vitro on primary murine HSCs as well as after systemic
administration in mice with healthy and fibrotic livers using intravital fluorescence microscopy. Coupling of NGFp to Ad via
S11 and/or PEGylation resulted in markedly reduced liver tropism and an enhanced adenoviral-mediated gene transfer to
HSCs. Transduction efficiency of both specific Ads was uniformly higher in fibrotic livers, whereas Ad.GFP-S11-NGFp
transduce activated HSCs better than Ad.GFP-PEG-NGFp. These experiments contribute to the development of a targeted
gene transfer system to specifically deliver antifibrotic compounds into activated HSCs by systemically applied adenoviral
vector modified with NGFp.
Kontakt
Institut für Experimentelle Gentherapie und Tumorforschung
Core-Facility Virale Vektor & Genom-Editing Technologien
Biomedizinisches Forschungszentrum
Schillingallee 69
D-18057 Rostock
Sekretariat
Ingrid Winkler
(+49) 381 494-5066(+49) 381 494-5062
ingrid.winkler@med.uni-rostock.de
Department Leben, Licht & Materie
Forschungsbau LL&M
Albert-Einstein-Str. 25
D-18059 Rostock
Forschungsbau LL&M
Albert-Einstein-Str. 25
D-18059 Rostock
