Polarized AAVR expression determines infectivity by AAV gene therapy vectors

Bradley A Hamilton; Xiaopeng Li; Alejandro A Pezzulo; Mahmoud H Abou Alaiwa; Joseph Zabner

doi:10.1038/s41434-019-0078-3

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Polarized AAVR expression determines infectivity by AAV gene therapy vectors

Journal article

Peer reviewed

Polarized AAVR expression determines infectivity by AAV gene therapy vectors

Bradley A Hamilton, Xiaopeng Li, Alejandro A Pezzulo, Mahmoud H Abou Alaiwa and Joseph Zabner

Gene therapy, Vol.26(6), pp.240-249

06/2019

DOI: 10.1038/s41434-019-0078-3

PMCID: PMC6588428

PMID: 30962536

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Abstract

Adeno-associated virus (AAV) has been investigated to transfer the cystic fibrosis transmembrane conductance regulator (CFTR) to airways. Inhaled AAV2-CFTR in people with cystic fibrosis (CF) is safe, but inefficient. In vitro, AAV2 transduction of human airway epithelia on the apical (luminal) side is inefficient, but efficient basolaterally. We previously selected AAV2.5T, a novel capsid that apically transduces CF human airway epithelia and efficiently restores CFTR function. We hypothesize the AAV receptor (AAVR) is basolaterally localized, and that AAV2.5T utilizes an alternative apical receptor. We found AAVR in human airway epithelia by western blot and RNA-Seq analyses. Using immunocytochemistry we did not find endogenous AAVR at membranes but overexpression localized AAVR to the basolateral membrane, where it preferentially increased transduction. Anti-AAVR antibodies blocked transduction by AAV2 from the basolateral side but not AAV2.5T from the apical side, suggesting a unique apical receptor. Finally, we found infection by AAV2 but not AAV2.5T was blocked by CRISPR knockout of AAVR in cell lines. Our data suggest the absence of apical AAVR is rate limiting for AAV2, and efficient transduction by AAV2.5T is accomplished using an AAVR independent pathway. Our findings inform the development of gene therapy for CF, and AAV vectors in general.

Cell Line

Cells, Cultured

Cystic Fibrosis Transmembrane Conductance Regulator - genetics

Cystic Fibrosis Transmembrane Conductance Regulator - metabolism

Dependovirus - genetics

Genetic Therapy - methods

Genetic Vectors - genetics

Humans

Receptors, Cell Surface - genetics

Receptors, Cell Surface - metabolism

Respiratory Mucosa - metabolism

Transfection - methods

Details

Title: Subtitle: Polarized AAVR expression determines infectivity by AAV gene therapy vectors
Creators: Bradley A Hamilton - University of Iowa
Xiaopeng Li - University of Iowa
Alejandro A Pezzulo - Roy J. and Lucille A. Carver College of Medicine
Mahmoud H Abou Alaiwa - University of Iowa
Joseph Zabner - University of Iowa
Resource Type: Journal article
Publication Details: Gene therapy, Vol.26(6), pp.240-249
DOI: 10.1038/s41434-019-0078-3
PMID: 30962536
PMCID: PMC6588428
NLM abbreviation: Gene Ther
ISSN: 0969-7128
eISSN: 1476-5462
Grant note: P30 DK054759 / NIDDK NIH HHS K01 HL140261 / NHLBI NIH HHS P01 HL051670 / NHLBI NIH HHS
Language: English
Date published: 06/2019
Academic Unit: Roy J. Carver Department of Biomedical Engineering; Pulmonary, Critical Care, and Occupational Medicine; Iowa Neuroscience Institute; Internal Medicine
Record Identifier: 9984359585202771

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