Molecular characterization of adeno-associated virus for the gene therapy of submucosal gland in cystic fibrosis / Dongsheng Duan.

Format:

Book

Manuscript

Microformat

Thesis/Dissertation

Author/Creator:

Duan, Dongsheng.

Contributor:

Engelhardt, John F., advisor.

University of Pennsylvania.

Language:

English

Subjects (All):

Penn dissertations--Pathology.

Pathology--Penn dissertations.

Pathology.

Academic Dissertations as Topic.

Medical Subjects:

Pathology.

Academic Dissertations as Topic.

Local Subjects:

Penn dissertations--Pathology.

Pathology--Penn dissertations.

Physical Description:

xiv, 244 pages : color illustrations ; 29 cm

Production:

1997.

Summary:

Cystic Fibrosis (CF) has emerged as a paradigm for the gene therapy of genetic disease. The full clinical potential of gene correction will only be achieved through a detailed understanding of both the target organ and the gene delivery vehicles. Dissection of the transduction process by recombinant adeno-associated virus (rAAV) suggested that this viral vector is an efficient and safe in vivo gene delivery system. Structural and functional examinations of integrated rAAV genomes have revealed head-to-head and head-to-tail genomes as the two predominant proviral forms. Importantly, only head-to-tail oriented proviral genomes were capable of being liberated upon challenge with Rep/Cap and adenovirus. These proviral structures led us to test the hypothesis that circularized genomes may also be an integral part of the AAV life cycle. Isolation of circular intermediates using a shuttle rAAV vector has confirmed this hypothesis and added to our understanding of AAV biology and rAAV mediated gene transfer. To better characterize appropriate progenitor cell targets for rAAV mediated gene delivery, we have evaluated the developmental aspects of airway submucosal glands. These studies have identified a transcription factor, lef-1, which was specifically expressed in submucosal gland progenitor cells prior to and during submucosal gland morphogenesis. The significance of the expression in gland progenitor cells was substantiated by in vivo antisense experiments. Inhibition of lef-1 expression resulted in an 8-fold decrease in both the number and the size of submucosal glands in a new-born ferret xenograft model. Furthermore, disruption of lef-1 in null mutant mice confirmed that this gene is necessary for nasal submucosal gland development. However, ectopic overexpression of lef-1 in airway xenografts using a bicistronic AAV-lef-1 GFP vector suggested that lef-1 was not sufficient for submucosal gland development. Transgenic mice which overexpress lef-1 under an airway specific promoter have further substantiated these hypotheses, indicating that lef-1 is necessary but not sufficient for gland development. Studies evaluating gene targeting to lef-1 expressing airway progenitor cells will lay the foundation for gene therapy of submucosal glands in CF.

Notes:

Supervisor: John F. Engelhardt.

Thesis (Ph.D. in Pathology) -- University of Pennsylvania, 1997.

Includes bibliographical references.

Local Notes:

University Microfilms order no.: 98-14837.

OCLC:

187456790