Gene transfer to airway epithelia in vivo with a feline immunodeficiency virus based lentiviral vector leads to long term expression of a beta-galactosidase transgene.

Gene Transfer for the Treatment of Inherited and Acquired Diseases
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The laboratory is performing a variety of gene transfer studies using integrating viral vectors from the retroviral family, especially lentiviruses derived from feline immunodeficiency virus (FIV). In addition, we are exploring the delivery of small interfering RNAs to respiratory epithelia to modify pulmonary disease states.

The goal of our gene transfer studies is to develop integrating vectors that could be applied to infants and children with cystic fibrosis and provide long-lasting treatment of lung disease. Recent studies have focused on FIV vector development, understanding barriers limiting transduction of airway epithelia, including cell proliferation and the localization of many retroviral receptors to basolateral membranes. Past and current approaches include:

1) transiently increasing epithelial proliferation (using growth factors such as KGF),
2) transiently disrupting epithelial junctions using calcium chelators, and
3) retargeting vectors by envelope pseudotyping.

The laboratory has developed novel FIV vector pseudotypes that target receptors on the apical surface of airway epithelia. These complementary strategies increase retroviral transduction efficiency to well differentiated epithelia.

In addition to the CF-related studies, the lab is broadly interested in the treatment of other inherited diseases with onset in early childhood, including hemophilia A. We are developing lentiviral gene transfer approaches for early intervention to prevent hemophilia A disease progression using a mouse model of factor VIII deficiency.

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Morphology of the apical surface of well-differentiated human airway epithelia. A, SEM view of the apical surface. A dense fibrous network was noted around cilia and microvilli on the apical surface (arrows). Spherical bodies suggestive of glycocalyceal bodies were also seen (arrow heads). B, ultra-thin sections of the airway epithelia were examined by transmission electron microscopy. The fuzzy apical glycocalyx around and in between the microvilli is indicated by the arrows. C, D) Components of the apical glycocalyx visualized using confocal microscopy. Apical surface staining for sialic acids.

Pseudotyping FIV lentivirus vector with the Ross river virus (RRV) glycoprotein improves mouse hepatocyte transduction in vivo following IV administration. A) vehicle alone, B) RRV pseudotyped FIV, C) VSV-G pseudotyped FIV.

Immunohistochemistry reveals expression of folate receptor alpha, a putative viral receptor, in polarized human airway epithelia.