title: Efficient Gene Delivery and Expression in Pancreas and Pancreatic Tumors by Capsid-Optimized AAV8 Vectors.
abstract: Despite efforts to use adeno-associated viral (AAV) vector-mediated gene therapy for treatment of pancreatic ductal adenocarcinoma (PDAC), transduction efficiency remains a limiting factor and thus improvement of AAV delivery would significantly facilitate the treatment of this malignancy. Site-directed mutagenesis of specific tyrosine (Y) residues to phenylalanine (F) on the surface of various AAV serotype capsids has been reported as a method for enhancing gene transfer efficiencies. In the present studies, we determine whether Y-to-F mutations could also enhance AAV8 gene transfer in the pancreas to facilitate gene therapy for PDAC. Three different Y-to-F mutant vectors (a single-mutant, Y733F; a double-mutant, Y447F+Y733F; and a triple-mutant, Y275F+Y447F+Y733F) and wild-type AAV8 (WT-AAV8) were administered by intraperitoneal or tail-vein routes to KrasG12D+/-, KrasG12D+/-/Pten+/-, and wild-type mice. The transduction efficiency of these vectors expressing the mCherry reporter gene was evaluated 2 weeks post administration in pancreas or PDAC and correlated with viral genome copy numbers. Our comparative and quantitative analyses of the transduction profiles demonstrated that the Y-to-F double-mutant exhibited the highest mCherry expression in pancreatic tissues (range 45-70%) compared with WT-AAV8 (7%; p < 0.01). We also detected a 7-fold higher level of vector genome copy numbers in normal pancreas following transduction with the double-mutant AAV8 compared with WT-AAV8 (10,285 vs. 1,500 vector copies/μg DNA respectively, p < 0.05). In addition, we observed that intraperitoneal injection of the double-mutant AAV8 led to a 15-fold enhanced transduction efficiency as compared to WT-AAV8 in mouse PDAC, with a corresponding ∼14-fold increase in vector genome copy numbers (26,575 vs. 2,165 copies/μg DNA respectively, p < 0.05). These findings indicate that the Y447+Y733F-AAV8 leads to a significant enhancement of transduction efficiency in both normal and malignant pancreatic tissues, suggesting the potential use of this vector in targeting pancreatic diseases in general, and PDAC in particular.
authors.affiliation: 1 Department of Anatomy and Cell Biology, University of Florida College of Medicine , Gainesville, Florida.
authors.forename: Min
authors.initials: M
authors.lastname: Chen
authors.affiliation: 1 Department of Anatomy and Cell Biology, University of Florida College of Medicine , Gainesville, Florida.
authors.forename: Kyungah
authors.initials: K
authors.lastname: Maeng
authors.affiliation: 1 Department of Anatomy and Cell Biology, University of Florida College of Medicine , Gainesville, Florida.
authors.forename: Akbar
authors.initials: A
authors.lastname: Nawab
authors.affiliation: 1 Department of Anatomy and Cell Biology, University of Florida College of Medicine , Gainesville, Florida.
authors.forename: Rony A
authors.initials: RA
authors.lastname: Francois
authors.affiliation: 1 Department of Anatomy and Cell Biology, University of Florida College of Medicine , Gainesville, Florida.
authors.forename: Julie K
authors.initials: JK
authors.lastname: Bray
authors.affiliation: 2 Department of Veterinary Medicine, University of Florida College of Medicine , Gainesville, Florida.
authors.forename: Mary K
authors.initials: MK
authors.lastname: Reinhard
authors.affiliation: 3 Department of Ophthalmology, University of Florida College of Medicine , Gainesville, Florida.
authors.forename: Sanford L
authors.initials: SL
authors.lastname: Boye
authors.affiliation: 3 Department of Ophthalmology, University of Florida College of Medicine , Gainesville, Florida.
authors.forename: William W
authors.initials: WW
authors.lastname: Hauswirth
authors.affiliation: 4 Department of Medicine, University of Florida College of Medicine , Gainesville, Florida.
authors.forename: Frederic J
authors.initials: FJ
authors.lastname: Kaye
authors.affiliation: 5 Division of Cellular and Molecular Therapy, Department of Pediatrics, University of Florida College of Medicine , Gainesville, Florida.
authors.forename: Georgiy
authors.initials: G
authors.lastname: Aslanidi
authors.affiliation: 5 Division of Cellular and Molecular Therapy, Department of Pediatrics, University of Florida College of Medicine , Gainesville, Florida.
authors.forename: Arun
authors.initials: A
authors.lastname: Srivastava
authors.affiliation: 1 Department of Anatomy and Cell Biology, University of Florida College of Medicine , Gainesville, Florida.
authors.forename: Maria
authors.initials: M
authors.lastname: Zajac-Kaye
mesh_terms: D000230:Adenocarcinoma / Q000235:genetics / Q000628:therapy*; D000818:Animals; D002213:Capsid; D021441:Carcinoma, Pancreatic Ductal / Q000235:genetics / Q000628:therapy*; D000229:Dependovirus / Q000235:genetics*; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics* / Q000627:therapeutic use; D006801:Humans; D051379:Mice; D016297:Mutagenesis, Site-Directed; D014161:Transduction, Genetic
keywords: AAV8; adeno-associated virus; gene therapy; pancreatic cancer